The Gordon Hammes ACS Biochemistry Lectureship
Professor Perry Frey, whose penetrating analyses of enzyme mechanisms have captivated the scientific community for more than 45 years, has been selected to present the second Gordon Hammes ACS Biochemistry Lecture at the 2010 ACS National meeting. The choice of Professor Frey recognizes his work on many problems, which has expanded the frontiers of the field of enzyme mechanisms.
Frey began his independent career by establishing the chemical mechanisms for the reactions catalyzed by uridinediphosphate galactose 4 epimerase (UDP-Gal epimerase) and galactose 1 phosphate uridylyltransferase. His work with the sugar epimerase provides an instructive example of the use of the substrate uridylyl group to anchor the reacting sugar at an enzyme active site where there are almost no specific sugar-protein interactions. This allows for oxidation of the sugar C-4 hydroxyl by a tightly bound NAD cofactor, rapid rotation of the 4-keto-sugar intermediate, and reduction of this intermediate to form epimerized product. Frey recognized early on that site-directed mutagenesis would revolutionize the study of enzyme mechanisms, and he used this as a tool in studies on both UDP-Gal epimerase and galactose 1-phosphate uridylyltransferase. He obtained X-ray crystal structures of wildtype and mutant enzymes in collaboration with Hazel Holden. This provided the structural information needed to draw mechanistic pictures for these two enzymatic reactions orders of magnitude more detailed than anything previously obtained from classical kinetic and structure-reactivity studies.
Frey was one of the leaders in the development of methods for the synthesis of nucleoside pyrophosphorothioates with chiral [18O]-labeled phosphorothioate groups, and in the use of these chiral compounds to determine the stereochemical course of enzyme-catalyzed thiophosphoryl transfers. Studies by Frey and several others were so successful that essentially all of the important mechanistic stereochemical problems relating to enzyme-catalyzed nucleophilic substitution reactions at tetravalent phosphorus were solved in less than fifteen years. During this time he also carried out important studies on the bond-order and charge delocalization at phosphorothioates.
Frey noted around 1994 an observation from 1972 by Robillard and Schulman that the proton bridging the δ nitrogen of His57 and the carboxyl group of Asp102 in the active site triad of chymotrypsin has an unusual downfield 1H-NMRresonance of 18 ppm. This anomalous chemical shift shows that the magnetic environment of the hydrogen-bonded active site triad proton is very different from that for hydrogen bonded protons in water, and is consistent with a much greater stability for this hydrogen bond compared to hydrogen bonds in aqueous solution. Frey followed his reevaluation of literature data with experimental work of his own design, that helped delineate the differences in the structure and stability of hydrogen bonds in water compared with hydrogen bonds in the highly organized and less polar environment of enzyme active sites. These thorough experiments and carefully argued conclusions have helped to draw the scientific community towards a consensus opinion about the contribution of hydrogen bonds to the rate acceleration for enzymatic reactions.
Frey’s studies on the bacterial enzyme lysine 2,3-aminomutase have provided critical insight into the mechanism of action of this member of the large radical SAM superfamily of enzymes. Lysine 2,3-aminomutase catalyzes the interchange of hydrogen and an amino group between adjacent carbon atoms, which is characteristic of adenosylcobalamin-dependent reactions. However, the protein catalyst does not contain or require this coenzyme and proceeded by a completely mysterious reaction mechanism. Frey has shown that the enzyme uses both S adenosylmethionine (SAM) and pyridoxal phosphate (PLP) as cofactors. The adenosyl group of SAM plays same mechanistic role as the B12-adenosyl group in the transport of hydrogen between substrate carbons. The α-pyridyl carbon of PLPwas shown to play a role similar to that of the cobalamine cobalt in mediating the skeletal rearrangement of substrate. Several hypothetical mechanisms for the generation of radicals by the reaction of SAM with the protein iron-sulfur center were considered, and strong evidence obtained to support a mechanism in which formation of a complex between the sulfonium sulfur of SAM and the iron-sulfur center is accompanied by an inner sphere electron transfer that occurs in concert with homolytic cleavage of the C-S bond in SAM.
Perry Frey has contributed heavily to the intellectual fabric of the community in which he has been a leader. His recently published book, Enzyme Reaction Mechanisms, will be the definitive text on this subject for the next twenty years, or longer. He has served as associate editor of Biochemistry since 1992, as co-chair GRC on Enzymes, Coenzymes and Metabolic Pathways and, as Chair of the organizing committee of the Winter Enzyme Mechanisms Conference. Professor Frey’s lecture at the 240th ACS National meeting in Boston (August, 2010) will be an important event in the history of Biochemistry and the Division of Biological Chemistry, who are joint sponsors of this lectureship.
SOURCE: American Chemical Society Division of Biological Chemistry