Research Topics
Department of Microbiology
Ph.D. (1968) Northwestern University
Research Topics
Research Interests
Protein degradation of bacteria functions to rid the cell of damaged or misfolded proteins and to generate amino acids under starvation conditions. In addition, proteolysis serves to regulate the levels of a number of important regulatory proteins. A complex proteolytic apparatus is required in order to carry out these reactions. The enzymes that carry out the initial steps in protein degradation produce peptides, typically 10-15 amino acids length, rather than free amino acids as products. These peptides are degraded by a group of peptidases that also participate in the catabolism of peptides supplied exogenously as nutrients. Our work is focused on understanding the nature of these peptidases and determining the specific role each enzyme plays in the degradation pathway. We are also interested in understanding how the structures of these enzymes determine their specialized functions in the peptide degradation process.
Several of the enzymes involved in peptide degradation have broad and overlapping specificities. One aspect of our recent work has involved an attempt to determine if these broad specificity enzymes have unique properties that allow them to carry out specialized functions. E.coli and S. typhimurium, the organisms we study, each contain two members of the leucineaminopeptidase family and both of these enzymes hydrolyze many of the same substrates. Only one of these enzymes, PepB, is able to hydrolyze peptides with N-terminal acidic residues and the other, PepA, also has special properties which allow it to attack certain peptides not hydrolyzed by other peptidases. We have shown that variation at a single amino acid position in Pep B determines its special affinity for N-terminal acidic peptides and have used this and other information to define signatures that we believe will allow definition of the specificities of new members of this peptidase family as they emerge from genome sequencing projects. We intend to pursue these studies to determine the unique contributions each of the peptidases makes to the overall degradation process.
In some cases, peptidases have very restricted specificities. Peptidase E for example, hydrolyzes only N-terminal aspartic dipeptides. We have been interested in understanding the structural basis for this unusual specificity. Our recently determined crystal structure for this enzyme shows that it is a member of a new class of serine hydrolase characterized by a unique Ser-His-Glu catalytic triad. We are pursuing further studies aimed at a detailed understanding of the structural basis for this enzyme's specificity. Several other enzymes in these organisms are highly specific for N-terminal aspartic peptides and we are interested in characterizing these enzymes, comparing them to Peptidase E, and determining their physiological role.
Key Words
Protein Purification and Characterization, Site Directed Mutagenesis, Microbial
Physiology, Genomics
Current Research Funding
NIH