regulatory strategies enzymes and hemoglobin. regulatory strategies what are the four ways in which...

Regulatory Strategies

Enzymes and Hemoglobin

Regulatory Strategies

What are the four ways in which enzymes are regulated?– allosteric regulation– enzymes exist in multiple forms (isoenzymes)– covalent modification– proteolytic cleavage

Allosteric Regulation

What are some of the characteristics of this form of regulation?– Activity influenced by non-covalent binding of

metabolite called a modulator• May be inhibitory or stimulatory

• May have one (monovalent) or several (polyvalent) modulators

• Binding induces shape change in enzyme

• Enzymes are large; two or more subunits

• Exhibit homotropic or heterotropic control


– enzyme may be part of a sequence in which the end product inhibits allosteric enzyme

– enzyme does not show Michaelis-Menten kinetics

Aspartate Transcarbamoylase

What is the reaction catalyzed by this enzyme?


CTP is a negative modulator and ATP is a positive modulator– feedback inhibition


How do we know that the catalytic and regulatory sites on this enzyme are distinct?– treat with

p-hydroxymercuribenzoate


Catalytic subunit consists of 3 chains (c3) and regulatory subunit consists of 2 chains (r2)


Allosteric modulators alter the quaternary structure of the enzyme


What is the difference between the T state and the R state of the enzyme?– T = tense; lower affinity for substrate– R = relaxed; higher affinity for substrate

Binding of substrate or substrate analog converts enzyme from T to R state– positive cooperativity– homotropic control


How does CTP act as a negative modulator?

Why is this an example of heterotropic modulation?


How does ATP act as a positive modulator?

Does this represent homo or heterotropic modulation?


Why is the mechanism just described called the concerted model?– All subunits must be

in same conformation, T or R


An alternative, the sequential model has been proposed


How does the sequential model differ from the concerted model?– Subunits may undergo individual sequential changes

in conformation

– Subunits can interact even in different conformations

– Change induced by binding of substrate to one subunits can increase or decrease substrate binding to other subunits

• positive or negative homotropic effects

• finer tuning

Allostery and Hemoglobin

Binding of O2 to hemoglobin represented by a sigmoidal curve similar to allosteric enzymes

Cooperativity promotes efficient O2 delivery


What part of the hemoglobin molecule binds O2?


How does the binding of O2 affect the structure of heme?


How does O2 binding influence the quaternary structure of hemoglobin?


How does 2,3 bisphosphoglycerate affect O2 affinity of hemoglobin?

How is this effect brought about?


How is fetal hemoglobin different from maternal hemoglobin?

How can this be explained?


What is the Bohr effect?What is the chemical basis of this effect?


CO2 also stabilizes deoxyhemoglobin by forming carbamate groups

Isozymes

What are isozymes or isoenzymes?– enzymes that have differences in amino acid

sequence but catalyze the same reaction• Example – lactate dehydrogenase

• Two different chains, M and H, exist

• Enzyme consists of 4 subunits– H4, H3M, H2M2, HM3, M4

• Each form has different Km and Vmax

Isozymes

M4 functions best in anaerobic environment while H4 in aerobic environment

Covalent Modification

Enzymes exist in active and inactive forms– Interconvertable by covalent modification

• Catalyzed by other enzymes

• Most modifications are reversible


What are the most common forms of covalent modification?– Phosphorylation and dephosphorylation

Which enzymes catalyze phosphorylation?– protein kinases

Which enzymes catalyze dephosphorylation?– protein phosphatases


What donates the phosphate group?


Why is phosphorylation an effective way to regulate proteins?– Phosphate group adds negative charges– Phosphate group can form hydrogen bonds– Free energy of phosphorylation is large– Can occur rapidly or slowly as needed– Can achieve amplification– Linked to energy status of cell


Example – glycogen phosphorylase– Two forms a and b

• A = active; 4 subunits each with a serine residue phosphorylated at OH group

• B = inactive; removal of PO4 groups causes protein to separate into two half molecules


What is the connection between c-AMP and protein kinases?– PKA activated by c-AMP in cells

Zymogens

What are zymogens?– inactive precursors of enzymes

How are they activated?– proteolytic cleavage

Zymogens

Zymogens

What are some other examples?– Blood clotting– Insulin– Conversion of procollagenase to collaginase in

metamorphosis– Conversion of procaspases to caspases in

apoptosis

Zymogens

Activation of chymotrypsinogen to chymotrypsin

Zymogens

Why is trypsin a key enzyme in zymogen activation?

Zymogens

Why do proteolytic enzymes have specific inhibitors?– To prevent premature activation of the enzyme– Activation of trypsin in pancreas could destroy

pancreatic tissue

Zymogens

How are zymogens involved in the formation of blood clots?

regulatory strategies enzymes and hemoglobin. regulatory strategies what are the four ways in which...

Documents

hemoglobin slide

enzyme slide

r slide

delivery slide

allosteric enzyme enzyme

heterotropic control

hemoglobin binding of

phydroxymercuribenzoate