Download MBBS Biochemistry PPT 49 Chemistry and Functions of Hemoproteins Chemistry Lecture Notes

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Chemistry and Functions of

Hemoproteins



Chemistry And Functions Of

Hemoglobin and Myoglobin
Synopsis

What are Hemoproteins?
What is Hemoglobin?
Structure of Hemoglobin
Functions of Hemoglobin
ODC and Factors affecting it
Normal Hb Variants
Hemoglobin Derivatives

INTRODUCTION
Hemoproteins

What Are Hemoproteins?

Hemoproteins are

Conjugated Proteins

With Heme as a

Prosthetic group in

their structures.
Hemoproteins are

Globular Proteins

(Whose Axial ratio less than 10)

Examples Of

HEME CONTAINING

PROTEINS AND ENZYMES

Of Human Body


Human Body Hemoproteins

Compounds with Heme group

Heme Containing Proteins
1. Hemoglobin (Hb)
2. Myoglobin (Mb)
3. Cytochromes (ETC Components)

Heme Containing Enzymes
1. Catalase
2. Peroxidase
3. Tryptophan Dioxygenase/

Tryptophan Pyrrolase

Hemoproteins are

vital for human body
Study Of Hemoglobin

What Is Hemoglobin?

Hemoglobin(Hb)

is a major

Hemoprotein of

Human body.


Hemoglobin Chemically is:

Conjugated Protein

In Hemoglobin

Heme is a Prosthetic group
Globin is a Protein part

(Hemoglobin = Heme + Globin)



Hemoglobin(Hb) is Red color

pigment

Location Of Hemoglobin-
Inside Red blood cells/Erythrocytes

of blood.


Amount Of Hemoglobin-

oEach RBC has approx

250-300 million Hb

molecules

oIn 25 x 1012 Erythrocytes

-750 gm of Hb
Hemoglobin In RBCs

Occupies:

33% of the RBC volume (1/3)

90-95% of the dry weight of

RBC is by Hb.

Normal concentration of

Hemoglobin in the Human Blood:

Adult Males-

13.5?17.5 gm/dL

Adult Females-

12.5?16.5 gm/dL
Hemoglobin Biosynthesis-

6.25 gm/day is the amount of Hb

biosynthesized during stages of

Erythropoiesis in bone marrow.

Synthesis of Hb begins in

Proerythroblast:

65% at Erythroblast stage

35% at Reticulocyte stage



Hemoglobin Function

Hb is associated to Respiration

Mechanism

Hb is a characteristic of Aerobic

life very important for survival.

Hb brings exchange of Gases-:
O2 and CO2

Terminologies of Hemoglobin

Hemoprotein -Heme is a prosthetic group
Chromoprotein - Red in color
Metalloprotein - Metal Iron (Fe) present
Respiratory Protein- Connected to

Respiration process and Respiratory

Chain(Electron Transport Chain)

Oxygen Binding Protein-Binds with

molecular Oxygen and transports it.
HISTORICAL ASPECTS

Of

Hemoglobin

Hemoglobin due to its

red color, has been of

interest since antiquity.
? Hemoglobin was a :

? First Protein to be crystallized -

1849.

? First Protein whose Mass

accurately measured.

? Mol weight of Hb-67,000

Daltons.

?First proteins to have X-ray

Diffraction structure

determined.

?First protein to be studied by

Ultracentrifugation.

?First protein to show that a

point mutation can cause

problems.
STRUCTURE OF HEMOGLOBIN

Two parts of Hemoglobin

Heme-Prosthetic group

Globin-Protein part
Hemoglobin Structural y

Tetrameric-contain 4 subunits.
Quaternary level of structural

organization.

Allosteric, Complex, Compact
Spheroidal= 64 x 55 x 50
Globular Protein

vHb of adults (Hb A) is a

Tetramer with

v4 Polypeptide subunits /4

Globin subunits

vConsisting of 2 - and 2 -

Globin chains
Each Subunit of Hemoglobin

contains:

1 Globin Chain and 1 Heme

group with a central Fe2+ ion

(Ferrous ion)

Hemoglobin Structure



Heme



One Hemoglobin molecule- 4 Subunits

One Subunit- 1 Polypeptide

Globin chain and 1 Heme moiety

Four Subunits- 4 Globin

Polypeptide chains+ 4 Heme

moieties.

1 Heme binds with 1 Oxygen molecule

4 Heme binds with 4 Oxygen molecule

1 OxyHb = 4 Globin+4Heme+4Oxygen


In Hb 4 polypeptide

chains are visualized as

two identical dimers,

()1 and ()2.


Two dimers are linked to

each other by weak polar

bonds- movement at the

interface of these two occurs

more freely.
Two polypeptide chains within a

dimer are held together tightly

by:

Ionic bonds and Hydrophobic

interactions, which prevent their

movement relative to each other.

v Thus Hb with

Quartenary structure is

in native conformation.


Significance of 4 Hb Subunits
11 and 2 2:

confirms stability of the

molecule.

1 2 and 2 1 :

confirms solubility of the

molecule.

1 2 and 1 2:
permit oxygenation and

deoxygenation.

v a2-b2 or a1-b1 interface has

35 amino acid residues

contact.

v a1-b2 and a2-b1 have 19

amino acid residue

contact.
Hemoglobin has

Outer relatively Hydrophillic

surface (Composed of polar a.a

/Provides Solubility)

Interior Hydrophobic (Made of

non polar a.a /insoluble a .a- Influences

Folding)

STRUCTURE /CHEMISTRY

OF HEME
Chem icWha

ally t

He I

m se H

is e

a me?

Ferroprotoporphyrin.

Prosthetic group of Hemoproteins
Red color pigment
Located interiorly in hydrophobic

Heme pocket present in Globin

subunit of Hb.

Metalloporphyrin


Chemically Heme is a

Ferroprotoporphyrin.
Heme Is- Ferroprotophoryin-IX

Protoporphyrin IX ring + Ferrous (Fe++)

Structure Of Heme
Structure Of Protoporphyrin IX-

Cyclic substituted Tetrapyrrole ring

structure (I,II,III and IV Tetrapyrrole ).

Tetrapyrrole rings has substituted

groups in systematic manner-

MV,MV,MP,PM

(M=Methyl ,V=Vinyl, P=Propionyl)

In Protoporphyrin ring ,Four

substituted Pyrrole rings are linked

by- 4 Methenyl bridges

Planar network of conjugated

double bonds of Heme absorbs

visible light and give red color

to Heme.


Structure of Heme
Iron in Heme

Functional form Iron in

Heme is-

Ferrous form(Fe++)
Reduced state

Fe++ located centrally

in Protoporphyrin ring

system.
Fe of Heme is

Hexavalent.

Fe of Heme forms 6 coordinated

bonds to satisfy its six valencies:

4 bonds linked with each Nitrogen

of 4 Pyrrole rings.

5th bond linked with Proximal

Histidine (F8) of Globin chain

( Globin=87, Globin=92).
6th bond is with Oxygen.
Binding of Iron in Hemoglobin-

Fe ++ is bound to :

4 Nitrogen of

Protoporphyrin ring

Globin chain ( Nitrogen of

Proximal His)

Oxygen

Iron content of Hb -

3.4 mg / gm of Hb


Role Of Iron in Hemoglobin

The Mineral, Iron, plays

indirectly an important

role in the body's

Delivery and use of

Oxygen by working

Muscles.



Iron helps in binding

Oxygen to Hemoglobin,

Oxygen get bound to Hb

then travels in the blood

stream to reach each and

every cell of the body.
Required amount of

Oxygen, delivery to

cells

Increases the body's

ability to perform

work.

Iron supports Aerobic Exercise

It has been postulated that a lack

of Iron in the body :

Reduces Aerobic capacity
Impair endurance

performance of exercise.


Iron of Heme gives red color

The visible absorption

spectra for Hemoglobin

The red color arises

from the differences

between the energy

levels of the d

orbitals around the

Ferrous atom.


Dietary Iron Deficiency

Features of Iron

deficient red blood

cells

Low number of

red blood cells

Hollow and

blanched red

cells

Iron deficiency is related to

Iron Deficiency Anemia

Low dietary Iron
Low Heme and Hb formation
Low Oxygen transport and release at

tissues and cells

Low cellular respiration
Low ETC operation in cells
Low ATP production in cells
Low cellular activity
Structure of Globin

Globin Subunits

Adult Hemoglobin has 4

Polypeptide chains

2 and 2 (identical pair).


Alpha Globin chains-

Composition- 141 amino acids
Molecular. Wt = 15,126 Daltons
Biosynthesis-Expression of

Globin gene on 16th Chromosome.

Beta Globin chains-

Composition- 146 amino acids
Molecular. Wt =15,866 Daltons
Biosynthesis- Expression of

Globin gene on 11th chromosome.


In Hemoglobin ?

2 (282 amino acid residues)
2 (292 amino acid residues)

Total 574 amino acids are

present in 1 Hemoglobin

molecule.
Each linear Globin

Polypeptide chain folds

To form 3 dimensional

Tertiary structure subunit.

Polypeptide chain has 8

Helices named as A, B, C, ...H.


Heme Pocket

Heme Pocket is a crevice/

hollow hydrophobic area

Formed in the interior of

Globin subunits

To locate the Heme moiety

in it.
The Heme pocket is surrounded

by E , F and H helices but not

with A , B, C ,D and G.

Heme group is tucked between
E and F helices of Globin subunit.

Amino acids in Globin chain are

identified by

The helix name and position

of a.a in that helix.

E7 His ( Distal His)
F8 His ( Proximal His)
Distal Histidine-E7 ( 58 , 63)

Proximal Histidine -F8 ( 87 , 92)

Fe++ of Heme is linked to

Proximal Histidine (F8)

O2 is linked to Distal

Histidine(E7).
Proximal and Distal His are

present in Heme pocket

In which Heme residue lies

and facilitates Oxygen binding.

Linking of Divalent O2 :

1.

Fe++ of Heme

2.

Nitrogen of Imidazole

group of Distal Histidine

of Globin chain( 58, 63)


Thus to attain stability

Oxygen is bound to

both Heme and Globin .



FUNCTIONS OF HEMOGLOBIN

Hemoglobin has important role in

Respiration mechanism-

v Hb Majorly Transports-

Oxygen (97% -100%)

v Hb Minorly Transports ?

Carbon dioxide (15% -25 %)
vDeoxy Hemoglobin

Transports-Protons(H+)

vThis is also termed as

Haldane effect

Hemoglobin Plays Role as Buffer-
(Hb/Hb-H+) in the Erythrocytes
Resists change in pH
Imidazole group of amino acid

Histidine of Hb molecule ?

Participates in buffering

mechanism of Hb.





Role Of Hemoglobin in Respiration

Since Hemoglobin has important role

in respiration mechanism, it is termed

as Respiratory Protein.

Respiratory Protein Hb serves in

transport and exchange of gases (O2

and CO2) between lungs and tissues.
How Significant Is

The Presence of Hemoglobin

To Human Body?

Why Natural y

There Is Presence

Of Hemoglobin

In the Living Bodies?
Presence of Hb in

blood

Gives less load to

Heart

Body cells requires approx. 500

gm/day of molecular Oxygen.

Molecular Oxygen is sparingly

soluble in body fluids.

This limits the Oxygen

transport in blood < 30 gm /day.


In fact if the body had to

depend upon dissolved

Oxygen in the plasma to

supply Oxygen to the cells.



The Heart would have to

pump 140 liters per minute.

Instead of normally 4 liters

per minute.

Hemoglobin a Polar, Oxygen

binding Protein/Oxygen

carrying Protein of blood.

Increases the binding and

effective transportation of

Oxygen through blood.
Presence of Hb in blood

facilitates the blood

To dissolve approx 70 times

more Oxygen than the plasma

without Hb can do.

oTotal Hb present in

each RBC

oCarry approx. More

than 1 billion Oxygen

molecules.
Thus to accomplish the following functions

Red blood cells has Hemoglobin (Hb):

Transfer of O2 from lungs to tissue

Transfer of CO2 from tissue to lungs

Hemoglobin serve as a

vehicle for transporting

the Oxygen

Through blood to reach

each and every cell.
Oxygen transported by Hb and

reached to every cell is used up in

Mitochondrial ETC

(Respiratory Chain/Cellular

respiration)

To generate ATP
(Oxidative Phosphorylation)

SALIENT FEATURES

OF

OXYGENATION

AND

DEOXYGENATION

OF HEMOGLOBIN
Oxygenation/Loading of

Oxygen

Hemoglobin gets Oxygenated

At Lungs
At increased pO2 concentration

(100-120 mm Hg)

At decreased pCO2

Saturation Of Oxygen By Hb
Normal ranges of pO2

100-120 mm Hg in arterial

blood at Lungs

35-40 mm Hg in venous

blood at tissues.

Hemoglobin is 97 % saturated

with Oxygen when it leaves the

Lungs-(Arterial Blood-Oxy Hb).

Under resting conditions Hb is

about 75% saturated with

Oxygen when it returns-(Venous

blood- Deoxy Hb).
Pulse Oximeter Is An

Instrument

That Measures The Percentage Hb

Ful y Saturated With Oxygen In

Arterial Blood


Thus the degree of

saturation with Oxygen is

related to:

Oxygen tension (pO2)
Oxygen requirement for

metabolic use at cellular

level

Features of Oxygenation of Hb



Oxygen binds with Hb to form HbO2

Oxygen links to Ferrous form

of Iron, of Heme

Non enzymatically, loosely

and reversibly.




During oxygenation One Hb

molecule with 4 Heme can

bind to four O2 molecules.

Binding of Oxygen to Heme of

Hb subunits:

Is weakly at low pO2

Is tightly at high pO2








Rate Of Hb Oxygenation:

? Less than 0.01 sec is required

for Hb Oxygenation.

?During Oxygenation

Ferrous of Heme is not

oxidized to Ferric.


v Oxygenation of

Hemoglobin causes

vConsiderable structural

conformational change

in Globin subunits.

Binding of Oxygen to Hb

rearranges the electronic

distribution and alters the d

orbital energy.
This causes a difference in

the absorption spectra.
Bluish for Deoxy Hb
Reddish for Oxy Hb
Measuring the absorption

at 578 nm allows an easy

method to determine the

percent of Oxygen bound to

Hemoglobin.






Ferric form of Iron is

non functional form

and cant bind with

Oxygen.

Deoxygenation/Unloading or

Offloading of Oxygen

Hemoglobin gets Deoxygenated

At Tissues
With Increased pCO2
Decreased pO2 levels (40 mm Hg)




















? Deoxy Hb has 2,3-Bis Phospho

Glycerate (23BPG) within it located

central y

? 2,3-BPG is pushed out of the Deoxy Hb

molecule during oxygenation

? Globin chains move closer when Hb

is Oxygenated.

Globin chains are pulled apart

when Hb is deoxygenated

This permits entry of 2,3-BPG

resulting in unloading of Oxygen



When Hb is fully

saturated with Oxygen

Each gram of

Hemoglobin is bound

with 1.34 ml of Oxygen.

COOPERATIVE BINDING

MECHANISM OF OXYGEN WITH

HEMOGLOBIN
Oxygen Binds To Hemoglobin

with

Cooperative Mechanism

Positive Al osteric Effect

Of Hemoglobin
Hb is an Allosteric Oxygen

binder with cooperative

mechanism.

Cooperative binding mechanism

is due to Tetrameric structure of

Hb.

Oxygen binding at the

four sites to the Heme

of Hemoglobin does not

happen simultaneously.
The binding of the first O2

to one subunit of Hb.

Enhances the binding of

futher O2 molecules to

remaining subunits of Hb

with greater affinities.

When Oxygen binds effectively

with one subunit

There increases the Oxygen

affinities for remaining

adjacent subunits, this is called

positive cooperativity.
Fourth Oxygen

molecule binds to fourth

subunit of Hb

300 times rapidly and

tightly as that of first

Oxygen bound to first

subunit.

Thus Hemoglobin is a

remarkable molecular

machine

That uses motion and

small structural changes

to regulate this action.
When a First Oxygen binds to Fe

in Heme of Hb,

The Heme Fe is drawn into the

plane of the Porphyrin ring.

This initiates a series of small

conformational changes that are

transmitted to adjacent Globin

subunits.

Oxygen ligand binding

information is

transmitted from one

subunit of Hb to another.
During Deoxygenation

Hemoglobin releases its bound

Oxygen.

As soon as the first Oxygen

molecule drops off, the

Hemoglobin starts changing its

shape.

This prompts the

remaining three Oxygen

molecules to be quickly

released.


In this positive

cooperative way

Hemoglobin picks up the

largest possible load of

Oxygen in the lungs,

And delivers the Oxygen

where and when needed.


T AND R FORMS

OF Hemoglobin
During loading and unloading

of Oxygen by Hb there occurs

considerable amount of

Allosteric movement.

This is due to the Oligomeric

/Tetrameric Structure of the

Hb molecule.

Models for Al osteric Behavior

Monod, Wyman, Changeux (MWC)

Model:

Allosteric Proteins can exist in two

states:

v R (Relaxed) State ? Oxy Hb
v T (Taut/Tensed) ? Deoxy Hb


Oxy & Deoxyhemoglobin


Quaternary structure of

Deoxy and Oxy Hemoglobin

T-state

R-state

The conformation of the Deoxy

state of Hb is called the T state

The conformation of the Oxy

state of Hb is called the R

state
T form of Hb

Deoxygenated Hemoglobin

is T form or Tensed/Taut

form of Hemoglobin

conformation.

T form /Tensed/Taut form of Hb has:

Centrally 2,3 BPG
Hydrogen instead of Oxygen
CO2

(Illustration Man with Three Tasks)

These moieties are held together by:

Eight salt bridges/ non covalent

interactions.

Thus T form is more constrained

form.
T form

predominates in

the absence of O2.

T form has lower

affinity for

Oxygen in low

pO2 environment.
At the center in T form of

Hb there occupies 2,3BPG

molecule which stabilizes

the Deoxy state of Hb.

Hb has more affinity for 2,3

BPG when pO2 is low.
Hence R form(OxyHb) at

low pO2

Gets attracted towards 2,3

BPG

Binds with it and looses its

Oxygen at Tissues.

R form of Hb

Oxygenated Hemoglobin

is a R form or relaxed

form of Hemoglobin

conformation.
During Oxygenation salt

bridges of T form are broken

T conformation is transformed

to R form.

R form is less constrained.

R form has

higher affinity for

Oxygen in high

pO2 environment.
At the Lungs where pO2 is

high

T form(Deoxy Hb) has now

higher affinity for O2, than 2,3

BPG

Hence T form binds with

Oxygen, extruding 2,3BPG

and get transformed to R

Form.

In R form of Hb

Only Oxygen is bound

R form has No linkage of

2,3BPG molecule
Protons
CO2
T R

Hb + pO2 HbO2

Deoxy-Hb Lungs Oxy-Hb

Increase of partial

pressure of Oxygen (pO2)

Causes the conversion of

T-form to R-form of Hb.




Transformation Of

T to R form of Hb is at Lungs
R to T form of Hb is at Tissues

Directly depends upon pO2

concentrations in the

environment of body and cells.
The conformational

changes of Hb from T to R

form and vice a versa are

known as "Respiratory

movement".

O2 binds much tighter to R

than to T.

R form of Hb(OxyHb) is more

negatively charged.


T-form (Tense/Taut)

has a much lower

oxygen affinity than

the R-form.

Oxy versus Deoxy Hemoglobin



Oxygenation

rotates the a1b1

dimer in

relation to a2b2

dimer about 15?




T Form of Hb

R Form Of Hb

Deoxy Hb is in T form

Oxy Hb is in R form binds

binds with CO2,H+ and

only with Oxygen

2,3BPG
T form has 8 salt bridges

Salt bridges are broken in

linked in between the dimer between the dimer subunits

subunits

during oxygenation of Hb.

More constrained form

Less constrained form

2,3 BPG is centrally located 2,3 BPG is extruded out from

in T form of Hb

R form of Hb

T form has low affinity for R form has higher affinity

Oxygen

for Oxygen

T form of Hb predominates R form of Hb predominates

in low pO2

at high pO2

Il ustration

Lungs ? Class Room
Tissues/Cells? House Environment
Oxygen- Study/Knowledge
Hemoglobin- Student
pO2 ?Teacher
Increased pO2- Knowledgeable and Skilled Teacher
Decreased pO2-Poor knowledge and Skill
T form of Hb- Student at House with

Dance,Sport,Internet

R form of Hb- Student at Class Room with Study
Oxygenation- Grasping of Knowledge
Deoxygenation- Revision /practice of Knowledge
Metabolic Condition-Examination
Significance of Tetrameric

Al osteric Structure

Hb being Tetrameric,

Allosteric protein facilitates

Cooperative binding

mechanism of Oxygen.

Enhances the efficiency of

Hb as an Oxygen transporter
Hb rapidly bind with oxygen

in lungs where pO2 is high (100

mm Hg)

Hb liberate Oxygen at tissue

capillaries where pO2 is low

(40mm Hg)

4 Factors Affecting

(Al osteric Effectors)

Loading and Unloading of Oxygen

At Lungs and Tissues
1. pO2 Concentration
2. pCO2 Concentration
3. pH (H+ Ion Concentration)
4. 2,3

BisPhosphoGlycerate

(2,3BPG/2,3DPG)

5. Glucose Concentration
6. Metabolic Condition

pO2 Concentration

At lungs pO2 concentration is

high pO2 =100-120 mmHg /torr

This favors oxygenation and

loading of oxygen by DeoxyHb.

DeoxyHb (T form) transformed to

form OxyHb (R form).
At Tissues pO2 concentration is low

35-40 mmHg /torr

This favors deoxygenation and

unloading of oxygen by OxyHb

OxyHb(R form) transformed to form

Deoxyhb (T form).

pCO2 And pH

At tissues due to active

metabolism

There is high

concentration of pCO2

and H+ ion concentration

(Low pH values).
Increased pCO2 and low pH at

tissues

Favors the OxyHb to loose

affinity for Oxygen,

Which in turn help in

unloading/off loading of oxygen

at tissues

(R form changes to T form).

The Bohr's Effect

Relation of Hemoglobin between

pCO2, pO2 and pH

Described by Danish Physiologist

Christian Bohr In 1904
The Bohr effect is a

physiological phenomenon

which states that:

Hemoglobin's Oxygen binding

affinity is inversely related for

both acidity and

concentration of Carbon

dioxide.

Thus The effect of

pCO2 and pH on

OxyHb is known as

Bohr's effect.
Bohr effect facilitates release of

Oxygen/Unloading Of Oxygen.



Since the tissues are relatively rich

in Carbon dioxide, the pH is

lower than in arterial blood;

Bohr effect is a

manifestation of

The acid-base equilibrium

of Hemoglobin.
CO2 + H2O CA H2CO3 CA H+ + HCO3-

Hydration of CO2 in tissues and

extremities leads to Proton production.

These Protons are taken up by Hb after

Oxygen released at tissues to Lungs.

The Protons transported by Hb are

released at the lungs.

Binding of protons to

Hb diminishes Oxygen

binding to Hb.

Binding of Oxygen to

Hb diminishes Proton

binding to Hb.
As the Proton(H+)

concentration increases


Affinity of Hemoglobin

towards Oxygen is

reduced.

At acidic pH (More H+ ion

concentration)-Favors unloading

of Oxygen from OxyHb

At alkaline pH(Less H+ ion

concentration)-Favors loading of

oxygen to Deoxyhb.
At lungs low pCO2 and low H+.
Favors oxygenation or loading of Hb

by O2.

Deoxyhb transports H+(protons)

from tissues to lungs.

On oxygenation of Deoxyhb, the

protons are liberated at lungs.

Effect Of 2,3BPG

on Loading and Offloading

Of Oxygen by Hb


2,3 Bis Phospho Glycerate

2,3 Bis Phospho Glycerate

(2,3BPG/2,3DPG) is an

intermediate of Rapaport

Leubering cycle

Related to Glycolysis inside

mature Erythrocytes.
2,3BPG is impermeable

to RBC membrane.

Glucose metabolism in

Erythrocytes increases the

concentration of 2,3BPG.

The "Inside" Story......

Where does 2,3-BPG bind ?

"Inside"
In the central cavity of Hb molecule.

What is special about 2,3-BPG ?

Negative charges interact with 2 Lys, 4

His, 2 N-termini of Globin.
At low pO2, 2,3BPG has high

affinity for adult Hb.

Increased 2,3BPG levels ?favors

Oxygen unloading by Hb.

Decreased 2,3BPG levels -favors

Oxygen loading by Hb.

The T form of Hb has 2,3

BPG centrally located

Which lowers the affinity

for Oxygen.
As the partial pressure of Oxygen

increases(pO2),

The 2,3, BPG is extruded out, and the

Hemoglobin resumes its original

state, known as the "Relaxed" or "R"

form,

R form has a high affinity for

Oxygen.

Conditions Of

High levels of 2,3BPG

During conditions of

cellular deprivation of

Oxygen.

2,3BPG levels in

Erythrocytes are increased
Conditions Of

High levels of 2,3BPG

Hypoxia
At high Altitudes
Severe Anemia
Lung Diseases
Cardiac disease -Anoxia
Blood loss

2,3 BPG levels in Hypoxia

2,3BPG levels increases in

hypoxia and at high altitudes.

Changes in 2,3-BPG levels

play an important role in

adaptation to hypoxia.
In hypoxic conditions pO2 is low

and

2,3 BPG levels are high


Due to affected metabolism of

Glucose in RBC's.

Increased 2,3-BPG levels

in red cells

Decreases Oxygen affinity
Facilitates unloading of

Oxygen to tissues.
Increased 2,3-BPG

also plays a role in

adaptation to

exercise.

Conditions of

Low 2,3BPG levels

Prolonged starvation
Erythrocyte disorders reduces

the levels of 2.3BPG.

Low 2,3 BPG reduces low

unloading of oxygen at tissue

level.
However formation of 2,3-BPG is

not very essential to life.

An individual who lacked the

enzymes necessary for 2,3-BPG

synthesis (Rapaport Leubering )

was perfectly well except for mild

Polycythemia.

The increased Oxygen

affinity of stored blood is

accounted (Blood Banks)

Due to reduced levels of

2,3-BPG.


Inosine addition to

stored blood in blood

bank

Increases the 2,3BPG

levels in it

This favors unloading

Oxygen on blood

transfusion.

OXYGEN DISSOCIATION CURVE

OF HEMOGLOBIN

(ODC)
Oxygen Dissociation Curve

(ODC) of Hemoglobin

ODC describes the relation

between

Partial pressure of Oxygen

(pO2) and percent saturation

of Oxygen by Hb.


ODC for Tetrameric ,Al osteric

Hb molecule is sigmoid shaped

(S shaped)



Sigmoid Shaped ODC Curve

Due To

Positive Allosteric Effect

Cooperative Binding Mechanism

Of Oxygen With Hb
p50 Of ODC

P5o is that pO2 value
Where the Hb is 50

percent saturated

with Oxygen.
P50 is 50% saturation

of Hb at pO2 of 27

mm Hg.

In ODC of Hb

P50 for Adult Hb

is 27 mm.Hg/torr
ODC depicts

O2 carrying

capacity of Hb

at different pO2

Salient Features Of ODC of Hb
Oxygen Dissociation

Curve Depicts:

Oxygen uptake and

release by Hemoglobin.

ODC Describes

The fractional saturation

of Heme groups of

Hemoglobin with Oxygen

at various Oxygen partial

pressures.
Normally the partial O2 pressure in the

Lungs is 100 mm.Hg and the Hb is

100 % saturated with O2.

In Tissues the partial oxygen pressure is

40mm.Hg and the Hb is 75% saturated

with O2.

100 - 75 = 25% of the O2 is released by

OxyHb and delivered to the tissues.

Percent Saturation Of Hb

At Different pO2

pO2 in torr

Percent Saturation

Of Hb

100 in Alveoli

97 %

40 in resting muscles

64 ? 75 %

20 in working muscles

20%

10 in vigorously exercising

10%

muscles
The sigmoid shape of the ODC curve

shows that:

With a small drop in partial O2

tension (pO2).

A significant amount of O2

release/offloading by OxyHb will

occur.

It is to be noted that the

OxyHb reaching to

tissues

Does not releases its

Oxygen completely at

one instance.
Instead the release of

Oxygen by OxyHb at

tissues is

As per the cellular need

for the Oxygen .

This regulated way of Oxygen

release by OxyHb at tissue level

May prevent from generation of

oxygen derived free radicals

(Reactive Oxygen Species: ROS)
Which in turn protect the

peroxidation of cellular

biomolecules by action of ROS.
FACTORS AFFECTING ODC

OR

ALLOSTERIC MODULATORS of ODC

The characteristics of normal

ODC depends upon following

factors:

Hemoglobin Structure


Environment within the

Erythrocyte
The environment Of

RBCs depends upon:

pO2
pCO2
pH(H+ concentration)
2,3-Bisphosphoglycerate
Glucose Concentration
Metabolic Condition
Temperature

Increased

H+,pCO2, 2,3BPG,Temperature

Causes unloading of Oxygen

from OxyHb.

They are Negative Allosteric

effectors of ODC.


Types And Conditions

Of ODC Shifts


Right Shift of ODC

With Its Conditions

If the Oxygen Dissociation

Curve is shifted towards Right

v Oxygen is unloaded by OxyHb



Oxygen affinity is decreased by Hb

Oxygen is not linked and

not retained in the Hb structure




















Conditions Which Shift ODC

To

Right Hand Side

Low Oxygen Affinity/Easy Oxygen Delivery/Easy

Unloading/ Prompt Offloading of Oxygen

? High pCO2 (Increased Metabolic States)

? High H+ (Acidosis)

? High 2,3-BPG: Hypoxic , Anoxic Conditions

? Exercise

? High body temperature : (Fever)

? Anemia : Hb S (low pO2)


Mnemonic for Factors causing

Right Shift of ODC :

CADET

C ? CO2
A ? Acid (H+)
D? 2,3-BPG /2,3 DPG
E ? Exercise
T ? Temperature

Left Shift of ODC

With Its Conditions
If the Oxygen Dissociation

Curve is shifted towards Left

Oxygen is not unloaded by OxyHb

Oxygen affinity is increased by Hb

Oxygen is linked and

retained in the Hb structure

Conditions Which Shift ODC

To

Left Hand Side














High affinity for Oxygen/ Low oxygen

Delivery/poor unloading of Oxygen

? High pO2

? In Alkalosis ( Low H+ high HCO3-)

? Low 2,3-BPG

? HbF

? Increased Methb and Carboxyhb

COMPARISON OF AN O2 DISSOCIATION CURVE AT NORMAL PH

AND WITH ACIDOSIS OR ALKALOSIS




Transport Of CO2 and H+

About 75 - 80% of tissue

Carbon dioxide is

processed and

transported in the form

of HCO -3(Bicarbonate ions)
Carbon dioxide

formed during

metabolism in tissues

is out in plasma

Then it diffuses freely

into the Erythrocytes

In aqueous solutions,

carbon dioxide undergoes a

pair of reactions

biocatalyzed by enzyme

Carbonic Anhydrase (CA).


Reaction 1

CO + H O H CO (Carbonic Acid)

2

2

2

3

Reaction 2

H CO H+ + HCO - (Bicarbonate ions)

2

3

3
Where the presence

of an enzyme

Carbonic

Anhydrase facilitates

reaction 1.



The H+ liberated in

reaction 2 are accepted by

deoxygenated

Hemoglobin, and

transported
The bicarbonate formed

in this sequence of reactions

Diffuses freely across the

red cell membrane and a

portion is exchanged with

plasma Cl-,

A phenomenon called the

"Chloride shift."

The bicarbonate ions are

carried in plasma to the

lungs


Where excretion of CO

2

occurs in the expired air.
Hb Minorly Transports CO2

15 ? 20% of CO2 is

Transported by Hb.
Transport of Carbon

dioxide by Hb, is unlike

that of Oxygen

CO2 does not bind to

Heme/Fe++ of Hb

CO2 is linked to

Globin part of Hb

and transported.
CO2 is bound to the

To Deoxygenated Hemoglobin
In Globin chains
At N-terminal Amino groups of
Valine residue
To form Carbaminohemoglobin

2 molecules of CO2 are

linked to 1 Hemoglobin

Transported through

blood from tissues to

Lungs and expired out.


? 5% of CO2 is

carried in free,

dissolved form

through blood.
Thus Deoxy Hb

carries:

CO2 and Protons

from Tissues to

Lungs.

At Lungs as Oxygen

gets bound to Deoxyhb

The CO2 and H+ comes

off of Deoxyhb and

expired out of Lungs.
At Lungs

At Tissue level

Respired air ?

Metabolism

pO2 is high ?90-100 mm Hg

pO2 is low-40mm Hg

pCO2 is high.

pH low (H+ high),2,3BPG high.

Hb is oxygenated to OxyHb (R Form)

OxyHb is dissociated to release

Cooperative binding mechanism of O2 to

oxygen at tissue level./O2 is

Hb

unloaded. OxyHb is deoxygenated

`T' form is transformed to `R' form.

R form is transformed to T form.

O2 binds to Fe ++ of Heme non

O2 released by Hb at tissue level is

enzymatically loosely and reversibly.

utilized for Biological Oxidation

and process(ETC).

4 O2 to 1 Hb

15-25% of Co2 is transported to

1.34 ml O2/gm of Hb transported

lungs by Hb forming

Carbaminohemoglobin and expired

out through lungs.

O2 is directly linked to Fe ++ of Heme and

CO2 is not linked to Fe ++ of Heme

distal His of 58 a.a and 63 a.a of Globin. But linked to amino groups of Val

residue.of Globin subunits

NORMAL HB VARIANTS
Normal Hb variants are

type of Hemoglobins

Present in different

physiological phases of

human life.

Role of Normal Hb Variant

:

To rightly fit for that

particular physiological

phase of life

Transport and Deliver

Oxygen as per need and

maintains normal cellular

activity.
Examples of Normal Hb Variants

Of Human Body

Globin Chain Synthesis,

starts at 3rd week of gestation.

Embryonic Stage/Embryonic

Hb -

vHemoglobin Gower I (z2e2)
vHemoglobin Gower II (a2e2)
vHemoglobin Portland (z2g2)
Fetal Stage:

v Major Hb : Hb F (a2g2)
v Minor Hb :HbA1(a2b2)

Adult Stage:

v Major Hb : Hb A1 (a2 b2)
v Minor Hb :

vHb A2 ( a2 d2)
vHb A3 (In old RBC's)

vHb F (a2 g2)
vGlycosylated Hb/Hb A1c
All Globin polypeptide chains are

homologous which arise from same

ancestral Genes.

Beta Polypeptide chain-146 a.a
Gamma chain-146 a.a
(differ in 39 a.a from chain )
Delta chain -146 a.a
(differ in 10 a.a from chain)

GLOBIN GENES

Of

Normal Hb Variants


Globin Gene Clusters


GLOBIN CHAINS In

Different Stages Of

Human life
Globin chain switch

Fetal Hemoglobin

(HbF)
HbF is a normal Hb

variant of fetal life

Hb F Predominates:

Fetus

New born infants

Fetal Hb (Hb F)

Globin part has : 2 and 2 subunits.
Globin chain differs

from Globin chain

in 39 amino acid

residues

Histidine residue at 143

position of Globin chain of

Hb A is replaced with

Serine a neutral amino

acid In Hb F .
Biosynthesis Of Hb F

Expression of following Globin genes

will produce and Globin chains

to form HbF:

Globin Gene located on 16 Chromosome
Globin Gene located on 11 Chromosome
Hb F biosynthesis starts by

7th week of gestation.

In Fetus Hb F predominates

during

Second and Third trimester

of gestation

At birth in newborn infants.

After birth there is rapid post

natal decline in HbF levels.

Within 4 months after birth

HbF is almost completely

replaced by Hb A.


Globin chain switch

Function and Features

Of HbF
HbF functions in

loading and unloading

of Oxygen in Fetus and

new born infants.

Hb F has a high

affinity for O2 than

HbA1.

Hb F has low

affinity for 2,3 BPG
HbF binds with O2 at

lower pO2 concentrations

than Hb A1.

P 50 for Hb F is 20 torr.

HbF has low Oxygen

releasing/unloading

capacity.

Thus ODC for HbF is

shifted towards Left.
Significance Of Hb F in Fetal Stage

The fetus is circulated

with maternal blood

Which has

comparative low pO2

as that of Lungs.
Hb F having high

Oxygen affinity

Gets oxygenated at low

pO2 of maternal blood.

This makes more

efficient trans placental

transfer of Oxygen from

maternal blood to fetal

HbF.
Thus Hb F in fetal phase rightly

fits for this state:

Since there is a low metabolic

activities in fetal cells and

requirement of low Oxygen.

Thus low release of Oxygen by

HbF suffice in this condition.

High levels of Hb F

In Adults Is Abnormal

Normally HbF in adults is less

than 1 %

HbF more than 1% in adult

hood is abnormal.
15-20% of HbF is found in patients

of Sickle Cell disease.

(Defect in Globin Genes)

More higher percent of HbF

is noted in individuals

suffering from

Thalassemia.

(Defect in Globin synthesis)
High levels of HbF in Adults will

have low release of Oxygen at

tissue levels:

Where the metabolic state and

requirement of Oxygen is high.

Thus HbF does not fit in adult

life.

P50 values for Hb A and HbF

P50 for Hb A= 27 mm.Hg
P50 for Hb F= 20 mm.Hg
















Adult Hemoglobins

Adult Hemoglobin Forms

Hb A1

Hb A2

Hb F

Globin chain a2b2

a2d2

a2g2

combinations

Normal %

96-98 %

1.5-3.2 %

0.5-0.8 %
Globin chain synthesis in adults

25% 25%

0.5% 1.5%

48%







25% 25%

0.5% 1.5% 48%

Chromosome 16

Chromosome 11

Hb A1
HbA1 is the major form

of Hb in adults and in

children over 7 months.

Globin has 2 and 2

subunits.

Hb A2
HbA2 has 2 and 2

Globin subunits.

Hb A2 is a minor form

of Hb in adults.

Hb A2 is 2 ? 3% of a total

adult Hb.

Hb A3
HbA3-altered form

of HbA1 found in

old RBC's.

Approx 3-10 %.

Hb A1

Hb F

P D

r iffe

edo

r

min eannc

t af etser be

1 yea tr w ofe Predominant in fetus and

birth and adults.

enn A

ew dult

born in - HbA

fants.

1

G and F

lobin ch e

a t

in al- HbF

-2 2

Globin chain-2 2

Less affinity towards O2 and More affinity towards O2

more affinity towards 2.3

and less affinity towards 2.3

BPG at tissues

BPG at tissues.

P50 is 27 mm.Hg

P5o 20 mm.Hg

Unloading power of oxygen Unloading power of oxygen

at tissue level is high.

at tissue level is low.

HbA1 denatured by alkali

Hb F resistant to alkali

denaturation.


ODC of Fetal Hb F vs. Adult Hb

GLYCOSYLATED

HEMOGLOBIN

(HbA1c)
Hb undergoes

spontaneous

glycosylation with

Glucose present in

Blood/RBCs.

The extent of glycosylation

with Hb depends on the

plasma concentration of

Glucose.


Once Hb is glycated

it remains till the

life span of RBC

(120 days).
Site Of Linkage Of Glucose

To Hemoglobin

Glucose is linked to Globin

part of Hemoglobin to Amino

acids:

Valine (terminal a.a ) of
Globin chain and
Lysine amino group


vLater the linked

Glucose is

transformed to

1-Deoxy Fructose.
Significance of Estimation

of

Blood Glycosylated Hemoglobin

Glycosylated Hb (HbA1c)

in normal healthy adults

is less than 5%

In Diabetes mellitus the

HbA1c is more than 5%
WHO Criteria for Diabetes Mellitus

HbA1c > 6.5%

Levels of Glycated Hb

gives idea of

Blood Glucose levels of a

person in last 3-4

months back.


Thus estimation of

Glycosylated Hb from blood

specimens in clinical

Biochemistry laboratory:

Gives Index of Glucose

Control in patients of known

Diabetes mellitus.


As the blood Glucose

levels increases

The percentage of

Glycosylated Hb increases
High Levels Of Glycosylated Hb

Decreases Oxygen Transport to

Tissues

Increased Glycosylated Hb

increases its affinity for

Oxygen.

Prevent release/unloading of

Oxygen at tissues

Induces hypoxia in extreme

cases
Increased Glycosylated Hb

Decreases Oxygen saturation

with Hb.

Increased Glycosylated Hb

Decreases Oxygen release at

tissues

Risk Of High Levels Of

Glycosylated Hb

In Patients Of Diabetes Mellitus
Diabetes mellitus Patients

Glycated hemoglobin of 6.5% -
Less risk for development of Diabetic

complications.

Glycated hemoglobin of 12 %-
High risk for development of Diabetic

complications.

HEMOGLOBIN DERIVATIVES
Hemoglobin interacts with chemical

agents to form Hb derivatives.

During formation of Hb derivatives

mostly Fe+2 part of Hb is involved.

Hemoglobin

O2

NORMAL HB DERIVATIVES
Normal Hb derivatives are

physiological and functional

forms of Hb.

Examples of Normal Hb derivatives.

OxyHb- Hb Bound to O2
Reduced Hb- Hb Bound to H+

ABNORMAL HB DERIVATIVES

or

Dyshemoglobins
Abnormal Hb Derivatives are

Acquired ones:

Abnormal Hb derivatives are

formed:

When blood interacts with

Chemical pollutants/Drugs

which has affinity for Hb.

Abnormal Hb derivatives has

Heme Iron linked to other

chemical compounds

instead of O2

OR

Hb is in a state where

Oxygen may not get linked

to Heme.
Examples of Abnormal Hb

Derivatives

1. Carboxyhemoglobin- CO linked to Fe+2 of Hb
2. Methemoglobin-Fe+2 of Heme transformed to

Fe+3

3. Cyanmethemoglobin-CN linked to Methb
4. Sulfhemoglobin-H2S interacted with Hb,
5. Sulfur linked to Fe +2 of Hb
6. Hematin- Ferriprotoporphyrin.
7. Hemin- Hematin Chloride.
8. Hemochromogen - Heme with denatured

Globin.

9. Cathemoglobin - Hematin with denatured

Globin.
Consequences of

Abnormal Hb Derivatives/

Dyshemoglobins

Dyshemoglobin Causes

Cyanosis

(Low Oxygen Saturation By Hb)
Dyshemoglobins in acquired states

affect normal structure and

function of Hb.

Dyshemoglobins are non

functional forms of Hb.

Dyshemoglobins affects Oxygen

transportation from lungs to

tissues.

CARBOXYHEMOGLOBIN

or

Carbon Monoxide Poisoning
Carbon Monoxide (CO)

is a colorless ,odorless,

toxic gas

Present in atmosphere as

chemical pollutant.

Sources of CO

Product of incomplete combustion of

fuel by vehicles.

Byproduct of Coal mines.
Cigarette Smoking (more than 4%).
Endogenous normal metabolism-

Heme catabolism(Heme Oxygenase

step)
CO has 200 times

more affinity for

Hb than O2.

CO readily links to Fe+2 of Hb and

form- Carboxyhb (Pink colour).

CarboxyHb has no place for binding

O2.

CarboxyHb reduces transportation

and delivery of O2 by Hb.


CarboxyHb delivers CO

at tissues instead of O2.

CO released in cells is

inhibitor of Cytochrome

oxidase in ETC.
CarboxyHb Toxicity

Toxicity due to CarboxyHb

is noted when concentration

is more than 20% in blood.

Concentration more than 40

-60% of Carboxyhb in body

may lead to death.
Symptoms of CarboxyHb Toxicity:

v Nausea

v Vomiting

v Headache

v Breathlessness

v Irritability

vFatigue

Investigation for CarboxyHb

Study of blood sample using

Hand Spectroscope.

Characteristic bands at 527

and 580 nm in green region

of visible spectrum confirms

presence of CarboxyHb.
Management and Treatment

Of CO Poisoning

Carbon monoxide poisoning

may be reverted

By increasing high

concentrations of O2
Cyanotic cases of Carboxyhb

treated by administration of

oxygen mask/Oxygen

cylinder.

Oxygen under high pressure

is helpful in managing

severe cases of CO toxicity.

Increased pO2 favors

replacement of CO by O2 to

form OxyHb transport and

deliver to tissues and support

the metabolic function.
METHEMOGLOBIN

Methemoglobin (MetHb) is an

abnormal Hemoglobin

derivative.

Methemoglobin has

Hematin/ Heme Iron in Ferric

(Fe+3) state.
Hematin is Ferriprotoporphyrin

Hematin +Globin = Methemoglobin

MetHb has defect in Heme

with normal Globin part.

MetHb has non functional

Iron- which cannot bind with

O2 and transport it.
Methemoglobin is non

functional oxidized

form of Hemoglobin.

Fe+3 of Hb gets

coordinated with water

instead of Oxygen at the

sixth position.
Formation of Methemoglobin

OR

Causes Of Methemoglobinemia

Normally about 1% of

Methemoglobin is

produced in blood

circulation.

Abnormal high levels of

blood Methemoglobin is-

Methemoglobinemia
Causes for Methemoglobinemia:

Acquired Cause :

Increases above 2% can

occur with the ingestion of

strong oxidant drugs

When blood is exposed to Oxidant Drugs

,Hb interacts with it and Fe+2 of Heme

truly gets oxidized to Fe+3.

Potassium Ferricyanide

Nitrites

Chlorates

Antipyrins

Sulfa Drugs (Sulfonamides)

Aniline Dyes
Conversion

Of

Methemoglobin

To

Hemoglobin

Reducing agents converts

Methemoglobin to functional Hb.

Glutathione

Ascorbic acid
Enzymes converts Methemoglobin

back to

Hemoglobin:

Methemoglobin Reductase
Cytochrome b5 Reductase

Conversion Of Methemoglobin To

Hemoglobin is

NADPH+H+ Dependent


H2O2 and Oxidant Drugs

Hemoglobin

Methemoglobin

(Fe+2)



(Fe+3)

Methemoglobin Reductase

NADP+ NADPH+ H+
The source of

NADPH+H+ for the use

of Methemoglobin

reductase activity is

Pentose Phosphate

Pathway(HMP Shunt).

Defect in HMP Shunt

affects the conversion of

Methemoglobin to

Hemoglobin due to devoid

of NADPH+H+


Congenital Causes of

Methemoglobinemia

As a result of deficiencies of

Methemoglobin

Reductase

G6PD enzyme of HMP

shunt
Familial Methemoglobinemia

Inherited deficiency of

Enzyme Methemoglobin

Reductase in the body

Causes Familial

Methemoglobinemia.

G6PD deficiency of HMP shunt

reduces generation of NADPH+H+

Which in turn affects

Methemoglobin Reductase

activity also leads to

Methemoglobinemia.
Methemoglobin Reductase in

absence of NADPH+H+

Does not convert

Methemoglobin back to

Hemoglobin.

Methemoglobin levels in blood

gradually increases to

Methemoglobinemia.

Consequences

Of

Methemoglobinemia
Methemoglobin is brown

colored pigment.

Hence

Methemoglobinemia ?

termed as :

Chocolate Cyanosis



Toxic Effects Of Methemoglobin

Methb has Fe+3 which is non

functional

Does not bind and transport

O2 to tissues.

Instead binds with water.
10-20% of Methemoglobin- Mild

Cyanosis.

50-60% of Methemoglobin- Severe

Cyanosis, Cardiopulmonary

Symptoms-Tachycardia, Depression.

More than 60% of Methemoglobin-

Unconsciousness and death.

Investigation Of MetHb

Study of Blood Sample

using Hand

Spectroscope.

Performing Schumm's

Test (Spectroscopy)
Management Of

Methemoglobinemia

Oral administration

of reducing agents

Ascorbic acid

Methylene Blue

Dried blood and old meat

have brown color.

Butchers uses Ascorbic acid

to reduce Methemoglobin to

make the meat look fresh!!
Sulfhemoglobin

Sulfhemoglobin - occurs when the

sulfur content of the blood

increases due to

Ingestion of sulfur containing

drugs

In chronic constipation( Gut

bacteria acts on unexcreted

material produces H2S)

Sulfhemoglobin is greenish

compound where sulfur is

covalently attached to

Porphyrin ring (Not to Iron).

Sulfhemoglobin cannot bind

with Oxygen.
Unlike the formation of

Carboxyhb and Methb,

The formation of Sulfhb is an

irreversible change of Hb.

Drugs producing Sulfhemoglobin:

Dapsone (Leprosy treating drug)
Phenacetin
Acetanilide
Sulfanilamides
These drugs produce Methb too
MYOGLOBIN

Myoglobin (Mb)-

Mb is a Hemoprotein of red

skeletal muscles.

Primarily occur in Cardiac

muscles.


Structure Of Myoglobin

Myoglobin is a

Monomeric unit

Mb is composed one

Globin chain and one

Heme moiety.

Myoglobin (Mb)
Mb is Spheroidal ,globular

molecule 44 x 44 x 25

Mb: Mol Wt 17,200 Daltons.

Myoglobin is rich in alpha

helix.

Globin part of Myoglobin is

composed of single

polypeptide chain


Composed of 153 amino

acid residues.
Myoglobin contains 1 Heme

group which binds with 1 O2 .

Iron in Mb is Fe2+ (Ferrous ion)

the functional form that binds

Oxygen.

Oxygen binds as the sixth

ligand to Fe (MbO2)

Myoglobin has very low

p50 value 2-3 torr/mm.Hg.

ODC of Myoglobin is

simple hyperbolic curve.


In comparison to Hemoglobin A and

HbF Myoglobin has high affinity for

Oxygen.

Hb F and Mb has low p50 values as

compared to p50 value of Hb A1.

With low p50 ,more Oxygen binds at

low pO2.
Function Of Myoglobin

Myoglobin is found in

cytosol of skeletal and

Heart muscles.
Myoglobin facilitates rapidly

Respiring muscle tissue

The rate of O2 diffusion from capillaries to

tissue is slow because of the solubility of

Oxygen.

Myoglobin increases the solubility of

Oxygen.

Myoglobin facilitates Oxygen diffusion.

Myoglobin in Muscle

Cells is a:

Oxygen storing

Hemoprotein

Reservoir of Oxygen
Myoglobin does not allow

Oxygen to remain in free state:



Oxygen diffused in muscle cells is

used up in Oxidative

phosphorylation.

If Oxygen remained unused in the

cells it immediately binds with Mb

to form MbO2

MbO2 releases/unloads

O2 when required.

MbO2 unloads oxygen

at extreme conditions

When pO2 of cellular

level reaches to 5 mm Hg.
Myoglobin releases

Oxygen in rapidly

respiring cells.

The released O2 is used

up in Oxidative

Phosphorylation.

Mb present within muscle cells comes

out in blood after damage to muscle

cells.

Mb is found abnormally in blood and

urine of MI cases.

Thus elevated Myoglobin levels in

blood/urine is a marker of

Myocardial damage.
Metmyoglobin does not

bind to Oxygen.

Since oxidation of Fe+2 yields

Fe +3 -Ferric iron

(non functional form)

Differentiate Between Hb and Mb
S.No Hemoglobin (Hb) Myoglobin (Mb)

1. Diffe

Hb rise

O nc

xyge e

n t s

ra nsof

port

Mb is Oxygen storing protein in

protein in RBCs of bloo Hb

d.

A

mu nd

scles. Mb

2.

Tetrameric has four Heme and Monomeric has one Heme and

binds with 4O2

binds with 1 O2.

3.

Oxygenated at Lungs

Oxygenated at Muscle Cell

Cytosol.

4.

HbO2 unloads oxygen at

MbO2 unloads oxygen at cell

tissues when pO2 is at 40

cytosol when pO2 is at 5 mmHg. To

mmHg.

rapidly respiring cells

P50 for HbA1 is 27 torr.

P50 for Mb is 2 torr.

5.

ODC is sigmoid shaped

ODC is hyperbolic shaped.

6.

Hb has 574 amino acids.

Mb has 153 amino acids.

Mol .wt-67,000 Daltons.

Mol wt-17,200 Daltons.

Cytochromes
Cytochromes ? Hemoprotein.

Cytochromes are components

of ETC

Who bring Oxidative

phosphorylation and

generates ATP .

Cytochrome P450-

Involved in Drug

detoxification.
Catalase and Peroxidases

Enzymes present richly in

Peroxisomes of cells.

Catalase and Peroxidase are of

Enzyme Class- Hydroperoxidases.



Glutathione

Peroxidase (R.B.C)

Leucocytes

Peroxidase (W.B.C)
Catalase and Peroxidase

Detoxify H2O2

Substrate for Catalase and

Peroxidase is H2O2 which detoxify

it.

Catalase and Peroxidase decomposes

2H2O2 to 2 H2O and O2.
Role of Catalase and Peroxidase

v Prevents accumulation of H2O2

(Toxic free radical) in cells .

vPrevents Peroxidation of membrane

lipids and protect cellular lysis.

Tryptophan Dioxygenase

Tryptophan Dioxygenase/Tryptophan

Pyrrolase involved in Tryptophan

catabolism.

Deficiency of Tryptophan Dioxygenase
Accumulates Tryptophan without its

breakdown to liberate Acetyl-CoA

(Ketogenic precursor) and Alanine

(Glucogenic precursor).
Deficiency of Tryptophan

Dioxygenase

Blocks Kynurenine

Pathway for the biosynthesis

of Niacin from Tryptophan.

Effect Of Cyanide and Carbon

Monoxide on Hemoproteins

CN and CO disrupts

physiological function of

HemoProteins.

Thus CN inhibits the function

of Hb ,Mb, Cytochromes.
Impaired activity of these

Hemoproteins

Badly affects Oxygen

metabolism and ATP

generation.

More affected cells are

Nervous system,

Erythrocytes.

Questions Of Hb Chemistry
1. Structure Of Hemoglobin
2. Heme Structure
3. Globin Structure
4. Functions of

Hemoglobin/Biomedical

Importance of Hemoglobin.

5. Salient features of Hemoglobin

Oxygenation and Deoxygenation.

6. Allosteric Effectors of Loading and

Unloading of Oxygen by Hemoglobin.

7. 2,3 BPG and its role in Hb.
8. ODC of Hemoglobin and factors

affecting it

9. CO2 Transportation in human body
10. Normal Hb Variants
11. Glycosylated Hb and its significance.
12. Hemoglobin Derivatives
13. Dyshemoglobins/Abnormal Hb

derivatives

14. CarboxyHemoglobin
15 . Methemoglobin
16. Types of Hemoproteins

Differentiate between following:
1. Hb A and Hb F
2. Hemoglobin and Myoglobin
3. T form and R form of Hb
4. Hb at Lungs and Hb at Tissues/
Oxygenation of Hb and

Deoxygenation of Hb.
THANKYOU

This post was last modified on 05 April 2022