Judith P. Klinman

Title
Professor Emeritus
Department
Dept of Molecular & Cell Biology
Phone
(510) 642-2668
Fax
(510) 643-6232
Research Expertise and Interest
catalytic & regulatory mechanisms in enzyme-catalyzed reactions, kinetic, spectroscopic, stereochemical biological techniques, peptide- derived cofactors, Nuclear tunneling & role of protein dynamics in catalysis, enzymatic activation of molecular oxygen
Research Description

The overall focus of our research is the determination of fundamental principles of catalytic and regulatory mechanism in enzyme-catalyzed reactions. Our experimental approach is broadly based and combines kinetic, spectroscopic, stereochemical, and molecular biological techniques.

Current Projects

Protein and peptide- derived cofactors. Work over the last decade has forced us to expand our definition of enzymatic cofactors to include structures that are derived from the protein side chains themselves. Early studies by others had implicated simple radicals (e.g., the tyrosyl and glycyl radicals), whereas more recent work, (including much from this laboratory) indicates complex structures that require novel chemical pathways. We are currently focused on four distinct structures that are derived from protein- or peptide-bound tyrosine side chains: the 2,4,5-trihydroxyphenylalanyl quinone (TPQ) found in the ubiquitous copper amine oxidases, the lysine tyrosyl quinone (LTQ) found in the mammalian lysyl oxidases, the cysteine tyrosyl radical (CTR) in the fungal galactose oxidase, and pyrroloquinoline quinone (PQQ), an essential vitamin for certain bacteria. Studies of the biogenesis of each of these structures require the expression of a precursor protein (or peptide in the case of PQQ), together with the establishment of conditions for monitoring the biogenetic processes. In the case of TPQ, LTQ and CTR all evidence indicates that these are produced by self-processing pathways that occur in the absence of exogenous protein factors, whereas PQQ requires six gene products. Both TPQ and LTQ were discovered in this laboratory and studies on the formation of TPQ are the most advanced, offering valuable protocols for studies of the other systems. In many instances, site specific mutagenesis has proven invaluable, leading to an accumulation of biogenesis intermediates that can be characterized by a variety of spectroscopic techniques. With regard to the function of these intriguing cofactors, recent studies of the mammalian TPQ-containing copper amine oxidases (found on the outer surface of the endothelium and adipocyte) suggest a role for enzymatically-produced hydrogen peroxide in cell signaling. Many experiments are either in progress or planned, to probe these provocative findings.

Copper- and iron-containing monooxygenases. A very large number of key biological functions involve the use of molecular oxygen. For example, dopamine beta-monooxygenase (DbM) catalyzes the formation of the hormone/neurotransmitter norepinephrine from dopamine. The enzyme is compartmentalized to either chromaffin vesicles in the adrenal gland or synaptic vesicles in the sympathetic nervous system. In contrast to amine oxidases, DbM contains only copper as a cofactor. Key mechanistic questions concern the role of copper in oxygen/substrate activation and the nature of reactive oxygen intermediates. We have developed a set of protocols to study O2 activation which includes distinguishing between O-16 and O-18 reactivity. We are now applying our unique methodology to a range of O2-dependent enzymes, which include dopamine and tyramine beta-monooxygenase; lipoxygenase (an iron protein), peptide amidation enzyme (a copper protein), tyrosine hydroxylase (a pterin/iron system, catalyzing dopa formation from tyrosine), glucose oxidase (a flavo-protein) and cytochrome P-450 (a heme/iron system). Patterns have begun to emerge regarding dioxygen reactivity. Our goal is to systematize biological systems with regard to their mechanism of dioxygen activation.

Nuclear tunneling in enzyme reactions. Over the course of our investigations of enzyme-catalyzed redox reactions, a number of anomalies had arisen which are incompatible with classical views of catalysis. In particular, these anomalies have suggested that quantum mechanical events may play a significant role in hydrogen transfer events at enzyme active sites. During the last several years, we have expanded the database for proteins that use tunneling as part of their catalytic strategy. Virtually every system examined thus far has shown some evidence for tunneling with the properties of tunneling varying depending on enzyme and protein structure, as well as temperature in the case of thermophilic enzymes. The effect of alteration in both substrate and protein structure is under investigation, with the goal of understanding the effect of changes in protein motion and active site geometry on tunneling. We have many interesting and surprising results that include the findings (i) that a single active site residue can control tunneling, and (ii) that thermophilic proteins utilize hydrogen tunneling even at the elevated temperatures of their optimal function.

In the News

October 3, 2014

Three faculty members awarded National Medal of Science

President Barack Obama has chosen three UC Berkeley faculty members – chemist Judith Klinman, applied mathematician Alexandre Chorin and the late statistician David Blackwelll – to receive the 2014 National Medal of Science. They were among 10 honorees announced Oct. 3 by the White House.

In the News

October 3, 2014

Three faculty members awarded National Medal of Science

President Barack Obama has chosen three UC Berkeley faculty members – chemist Judith Klinman, applied mathematician Alexandre Chorin and the late statistician David Blackwelll – to receive the 2014 National Medal of Science. They were among 10 honorees announced Oct. 3 by the White House.

Loading Class list ...
.