This project is focused on understanding the physical and mechanistic properties of enzymes that underlie their exquisite function. In recent years, protein motions have been implicated as essential to achieve an extremely rapid catalysis of bond cleavage events. A spatial and temporal resolution of such protein motions is being pursued using enzyme prototypes that catalyze hydrogen and methyl transfer reactions. A description of the hierarchy and range of the requisite protein motions (covering time scales that can differ by ca. 1015 fold) will take place utilizing highly developed kinetic and biophysical probes. A second emerging area in biological catalysis concerns the post-translational modification of peptides that have been synthesized at the ribosome. A combination of structural and biochemical probes is addressing the enigmatic pathway that produces the bacterial cofactor and vitamin, pyrroloquinoline quinone. As the result of a number of recent breakthrough observations, it appears that resolution of this long-standing problem is now within reach.

Public Health Relevance

We are studying the physical and mechanistic principles that underlie protein function. The coupling of enzyme motions to the chemical steps of catalysis is being interrogated, as are novel enzyme activities that produce ribosomally-synthesized, post-translationally modified products.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Unknown (R35)
Project #
5R35GM118117-05
Application #
9892015
Study Section
Special Emphasis Panel (ZGM1)
Program Officer
Barski, Oleg
Project Start
2016-04-01
Project End
2021-03-31
Budget Start
2020-04-01
Budget End
2021-03-31
Support Year
5
Fiscal Year
2020
Total Cost
Indirect Cost
Name
University of California Berkeley
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
124726725
City
Berkeley
State
CA
Country
United States
Zip Code
94710
Offenbacher, Adam R; Iavarone, Anthony T; Klinman, Judith P (2018) Hydrogen-deuterium exchange reveals long-range dynamical allostery in soybean lipoxygenase. J Biol Chem 293:1138-1148
Zhu, Wen; Martins, Ana M; Klinman, Judith P (2018) Methods for Expression, Purification, and Characterization of PqqE, a Radical SAM Enzyme in the PQQ Biosynthetic Pathway. Methods Enzymol 606:389-420
Klinman, Judith P; Offenbacher, Adam R (2018) Understanding Biological Hydrogen Transfer Through the Lens of Temperature Dependent Kinetic Isotope Effects. Acc Chem Res 51:1966-1974
Vaughn, Morgan B; Zhang, Jianyu; Spiro, Thomas G et al. (2018) Activity-Related Microsecond Dynamics Revealed by Temperature-Jump Förster Resonance Energy Transfer Measurements on Thermophilic Alcohol Dehydrogenase. J Am Chem Soc 140:900-903
Barr, Ian; Stich, Troy A; Gizzi, Anthony S et al. (2018) X-ray and EPR Characterization of the Auxiliary Fe-S Clusters in the Radical SAM Enzyme PqqE. Biochemistry 57:1306-1315
Tu, Xiongying; Latham, John A; Klema, Valerie J et al. (2017) Crystal structures reveal metal-binding plasticity at the metallo-?-lactamase active site of PqqB from Pseudomonas putida. J Biol Inorg Chem 22:1089-1097
Horitani, Masaki; Offenbacher, Adam R; Carr, Cody A Marcus et al. (2017) 13C ENDOR Spectroscopy of Lipoxygenase-Substrate Complexes Reveals the Structural Basis for C-H Activation by Tunneling. J Am Chem Soc 139:1984-1997
Hu, Shenshen; Soudackov, Alexander V; Hammes-Schiffer, Sharon et al. (2017) Enhanced Rigidification within a Double Mutant of Soybean Lipoxygenase Provides Experimental Support for Vibronically Nonadiabatic Proton-Coupled Electron Transfer Models. ACS Catal 7:3569-3574
Latham, John A; Barr, Ian; Klinman, Judith P (2017) At the confluence of ribosomally synthesized peptide modification and radical S-adenosylmethionine (SAM) enzymology. J Biol Chem 292:16397-16405
Evans 3rd, Robert L; Latham, John A; Xia, Youlin et al. (2017) Nuclear Magnetic Resonance Structure and Binding Studies of PqqD, a Chaperone Required in the Biosynthesis of the Bacterial Dehydrogenase Cofactor Pyrroloquinoline Quinone. Biochemistry 56:2735-2746

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