Documents about Nucleophilic Attack

  • 9 Pages

    Lecture 18a

    Cornell, BIOBM 3310

    Excerpt: ... LL PRECISELY ALIGNED. THIS PRECISE ALIGNMENT IS BUILT INTO THE ENZYME STRUCTURE. HIS57 ACTS AS BASE; SIMULTANEOUSLY, SER195-O MAKES NUCLEOPHILIC ATTACK ON CARBONYL C OF PEPTIDE BOND THIS STEP IS RATEDETERMINING FOR PEPTIDE HYDROLYSIS i.e. TRANSITION STATE IS HERE! ES OXYANION "HOLE": 1. GOOD STERIC FIT TO O , BUT ONLY IN ITS Asp 194 TETRAHEDRAL ORIENTATION 2. H-BONDS FORM TO PEPTIDE N-H OF GLY193 195 O ATTACK AND SER NOT THE TS, BUT NOT SO STABLE ES' CHYMOTRYPSIN MECHANISM NOW HIS57 ACTS AS AN ACID ELECTRON DENSITY FLOWS OUT OF THE C N PEPTIDE BOND H THE FIRST PRODUCT, A PEPTIDE FRAGMENT, LEAVES THE ENZYME AND DIFFUSES AWAY H ES' NOW INTEMEDIATE IS BOUND WITH SOME STABILITY TO CHYMOTRYPSIN THIS COVALENT INTERMEDIATE HAS BEEN CLEARLY IDENTIFIED: TRAPPED IN CRYSTALS OF CHYMOTRYPSIN AT -55oC, REVEALED BY XRAY DIFFRACTION ES' CHYMOTRYPSIN MECHANISM WATER BINDS IN THE ACTIVE SITE WATER REPLACES FIRST PEPTIDE FRAGMENT IN ACTIVE SITE NOT BULK WATER! A SINGLE H2O, BOU ...

  • 3 Pages

    Lecture 18

    Cornell, BIOBM 3310

    Excerpt: ... are donated/accepted, or an electron pair donated/accepted, but is in contrast to "specific" acid or base catalysis, in which the catalyst is the H+ ion or the OH- ion). The positive charge on this His+ is stabilized by Asp-102. [Note the catalytic triad of His, Ser, Asp] Simultaneous with the transfer of the proton to the His, the Ser-O: makes a nucleophilic attack on the carbonyl C of the peptide bond. This nucleophilic attack leads to the intermediate ES' shown after step 2. This ES' tetrahedral intermediate is not the transition state, but the TS is "something like" ES'. In fact, the actual TS for this overall rxn is indeed believed to occur just prior to formation of ES', and the actual TS is believed to have many of the properties of this tetrahedral intermediate. This tetrahedral intermediate ES' fits into a pocket in the enzyme that fits the oxygen anion only when it is tetrahedral to the peptide carbonyl C. This is the enzymes "oxyanion hole." It makes both a steric and a chemical fit between t ...

  • 4 Pages

    Lecture 19

    Cornell, BIOBM 3310

    Excerpt: ... xt intermediate. p. 132 For the family of enzymes known as the Serine proteases, here is the overall reaction equation for the catalyzed reaction: E +S ES ES' ES' P1 ES' EP2E + P2 The rate of enhancement over the uncatalyzed hydrolysis of peptide is about 10 10. What if we mutate either Ser195 or His57? You might imagine that catalysis would be completely ruined. Indeed, the rate enhancement drops to about 10 5. This is a big drop of about 100,000-fold, but there is still impressive catalytic power left! Catalytic power still comes from the binding, hence stabilization, of the tetrahedral transition state. A summary of some key catalytic features of chymotrypsin: Ser195 -O: binds to the peptide carbonyl C ( nucleophilic attack ) Peptide NH of Ser195 and Gly193 H-bond to the O- of the tetrahedral transition state (and tetrahedral intermediates) Enzyme shape fits the tetrahedral intermediate, thereby especially lowering TS free energy. In other words, the enzyme is specialized to fit the TS especi ...