Seminars this week for credit:

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Seminars this week for credit

• Dec 04 M
• Clint Spiegel, Ph.D.
• University of California, Santa Cruz, "Ribosome
  Function in an RNA World
• 400 p.m. SL 110
• Peggy Daley

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Final Exam W December 13 1030 am
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http//video.nbc.com/player.html?dlid45139
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Chapter 15 Catalytic Mechanisms
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Catalytic Mechanism Determination
1) kinetic analysis - what is the kinetic
signature? - mode of inhibition revealed
- determine rates for individual steps - does
order of addition matter? (sequential vs.
Ping-Pong) 2) active site modification
(irreversible inhibitors) - derivatize protein
identify the modified sidechain(s) 3) structure
determination (e.g. RNase A, lysozyme, serine
proteases)
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Enzymatic catalysis proceeds by one or more of
1) general acid/base catalysis (GABC) 2)
covalent catalysis 3) electrostatic
stabilization 4) proximity effects 5)
preferential stabilization of the
Effects 1 and 3 are often manifested as effect
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• http//cti.itc.virginia.edu/cmg/Demo/mechanism/me
  ch.html
• Chymotrypsin
• ser protease

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Covalent catalysis
Is characterized by the formation of a covalent
Enz-S adduct that alters the reaction
pathway Nucleophiles many amino acid
sidechains (H, K, C, S, D, E, Y), some cofactors
(TPP) Electrophiles some cofactors (e.g. PLP)
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Stryer Fig. 9.2 Identifying an active ser--Out
of 28 ser, only 195 is labeled
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Figure 15-19 Reaction of TPCK with chymotrypsin
to alkylate His 57.
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Stryer Fig. 9.3 Chromogenic substrate
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Figure 15-18 Time course of p-nitrophenylacetate
hydrolysis as catalyzed by two different
concentrations of chymotrypsin.
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Stryer Fig. 9.5 Covalent catalysis
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Figure 15-21 The active site residues of
chymotrypsin.
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Figure 15-23 Catalytic mechanism ofthe serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-23 Catalytic mechanism of the serine
proteases.
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Figure 15-25a Transition state stabilization in
the serine proteases. (a) The Michaelis complex.
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Figure 15-25b Transition state stabilization in
the serine proteases. (b) The tetrahedral
intermediate.
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Table 15-4 A Selection of Serine Proteases.
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Figure 15-22 Relative positions of the active
site residues in subtilisin, chymotrypsin, serine
carboxypeptidase II, and ClpP protease.
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(No Transcript)
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Example of convergent evolution.
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Stryer Fig. 9.16 Site directed mutagenesis of
subtilisin. Note the log scale. Mutations in
the catalytic triad lead to a dramatic loss of
activity
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Figure 15-24a TrypsinBPTI complex. (a) The
X-ray structure shown as a cutaway surface
drawing indicating how trypsin (red) binds BPTI
(green).
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Figure 15-3 The bovine pancreatic RNase
Acatalyzed hydrolysis of RNA is a two-step
process with the intermediate formation of a
2,3 -cyclic nucleotide.
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Figure 15-2 The pH dependence of Vmax/KM in the
RNase Acatalyzed hydrolysis of cytidine-2,3
-cyclic phosphate.
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