Chymotrypsin

While catalysis by aspartic proteases involves the direct hydrolytic attack of water on a peptide bond, catalysis

Ala ^CHO /CH2NH2 PJr KG /CH2NH2 ^CHO Gu

E — CHO ^ > E ->- E -— CH2NH2 ^ > E ->■ E -E — CHO

Figure 7-4. Ping-pong mechanism for transamination. E—CHO and E—CH2NH2 represent the enzyme-pyridoxal phosphate and enzyme-pyridoxamine complexes, respectively. (Ala, alanine; Pyr, pyruvate; KG, a-ketoglutarate; Glu, glutamate).

Figure 7-5. Two-dimensional representation of Koshland's induced fit model of the active site of a lyase. Binding of the substrate A—B induces conformational changes in the enzyme that aligns catalytic residues which participate in catalysis and strains the bond between A and B, facilitating its cleavage.

Figure 7-5. Two-dimensional representation of Koshland's induced fit model of the active site of a lyase. Binding of the substrate A—B induces conformational changes in the enzyme that aligns catalytic residues which participate in catalysis and strains the bond between A and B, facilitating its cleavage.

by the serine protease chymotrypsin involves prior formation of a covalent acyl enzyme intermediate. A highly reactive seryl residue, serine 195, participates in a charge-relay network with histidine 57 and aspartate 102. Far apart in primary structure, in the active site these residues are within bond-forming distance of one another. Aligned in the order Asp 102-His 57-Ser 195, they constitute a "charge-relay network" that functions as a "proton shuttle."

Binding of substrate initiates proton shifts that in effect transfer the hydroxyl proton of Ser 195 to Asp 102 (Figure 7-7). The enhanced nucleophilicity of the seryl oxygen facilitates its attack on the carbonyl carbon of the peptide bond of the substrate, forming a covalent acyl-enzyme intermediate. The hydrogen on Asp 102 then shuttles through His 57 to the amino group liberated when the peptide bond is cleaved. The portion of the original peptide with a free amino group then leaves the active site and is replaced by a water molecule. The charge-relay network now activates the water molecule by withdrawing a proton through His 57 to Asp 102. The resulting hydroxide ion attacks the acyl-enzyme in-

H Hh

AspY

Asp X

Asp Y

Asp X

CH, AspY

CI O OCO

Asp X

Figure 7-6. Mechanism for catalysis by an aspartic protease such as HIV protease. Curved arrows indicate directions of electron movement. CD Aspartate X acts as a base to activate a water molecule by abstracting a proton. CD The activated water molecule attacks the peptide bond, forming a transient tetrahedral intermediate. CD Aspartate Y acts as an acid to facilitate breakdown of the tetrahedral intermediate and release of the split products by donating a proton to the newly formed amino group. Subsequent shuttling of the proton on Asp X to Asp Y restores the protease to its initial state.

Asp 102

Ser195

His 57

Asp 102

Ser195

His 57

Ser195

Asp 102

His 57

Asp 102

Ser195

His 57

Asp 102

Ser195

His 57

Ser195

His 57

Asp 102

Figure 7-7. Catalysis by chymotrypsin. CD The charge-relay system removes a proton from Ser 195, making it a stronger nucleophile. CD Activated Ser 195 attacks the peptide bond, forming a transient tetrahedral intermediate. CD Release of the amino terminal peptide is facilitated by donation of a proton to the newly formed amino group by His 57 of the charge-relay system, yielding an acyl-Ser 195 intermediate. CD His 57 and Asp 102 collaborate to activate a water molecule, which attacks the acyl-Ser 195, forming a second tetrahedral intermediate. CD The charge-relay system donates a proton to Ser 195, facilitating breakdown of tetrahedral intermediate to release the carboxyl terminal peptide ©.

termediate and a reverse proton shuttle returns a proton to Ser 195, restoring its original state. While modified during the process of catalysis, chymotrypsin emerges unchanged on completion of the reaction. Trypsin and elastase employ a similar catalytic mechanism, but the numbers of the residues in their Ser-His-Asp proton shuttles differ.

Diabetes 2

Diabetes 2

Diabetes is a disease that affects the way your body uses food. Normally, your body converts sugars, starches and other foods into a form of sugar called glucose. Your body uses glucose for fuel. The cells receive the glucose through the bloodstream. They then use insulin a hormone made by the pancreas to absorb the glucose, convert it into energy, and either use it or store it for later use. Learn more...

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