Expert in Protein Chemistry & Regulation, Proteomics, Protein Mass Spectrometry, Proteases, Enzymology
As an investigator, Expert has studied structure-function relationships among enzymes for many years. The subjects have included proteases, protein kinases, phosphatases, proteases, and phosphodiesterases. These studies have included probes of active sites with chemical reagents, studies of the kinetics and the inhibition of enzymes, and studies of regulatory mechanisms such as phosphorylation, allostery, and zymogen activation.
As an investigator, Expert has spent more than 30 years determining the amino acid sequences of proteins using classical methods of proteolytic cleavage, chromatographic separations, Edman degradations, and mass spectrometry.
From 1959 to about 1980, Expert studied a wide range of proteases including all those found in the pancreas, many found in blood coagulation, and some found in fertilization and in secretory mechanisms. These studies included determination of their structures and examination of the relationship between the structure of each protease, its function, and its inhibitors.
Expert has expertise in protein mass spectrometry using matrix-assisted laser desorption ionization-time of flight analysis (MALDI-TOF). This technique is found to complement electrospray instrumentation for examining the molecular weight of proteins and peptides in mixtures, identifying proteins, identifying their posttranslational modifications.
Expert is familiar with degradative techniques for determining amino acid sequence (Edman egradation, enzymatic digestions); chemical modification of proteins to alter stability or activity; and purification of proteins and peptides.
Expert has studied proteases and their zymogens, particularly trypsin and other serine proteases. He is interested in activation, activity, specificity, mechanism, homology, biological roles, etc.
In the course of determining the amino acid sequence of a wide range of proteins over a 25 year period, Expert has dealt with a whole range of posttranslational modifications. These include phosphorylation, glycosylation, gamma carboxylation, acetylation, methylation, biotinylation, and myristylation. Recently the ESI-MS and MALDI-TOF technologies has proven to be a particularly valuable tool for such studies.
Expert has focused upon mechanisms of limited proteolysis that convert inactive zymogens to active enzymes in systems as diverse as the digestive and blood coagulation systems. In addition, he has used limited proteolysis as an experimental probe for domain boundaries.
Expert’s laboratory focused on analyzing amino acid sequences and improving the efficiency of those analyses from 1959-1990. Bovine trypsin was sequenced in 1964 (223 residues). 22 years (and many proteins) later human von Willebrand Factor (2050 amino acids) was sequenced. The lab built one of the first automatic sequencers in the U.S., which contributed to the development of the first commercially available sequencer. These studies provided early indications of the homologous relationships among proteins and of the domain nature of their construction.
An obvious question about a protease like trypsin or thrombin is why it does not digest the proteins found in the cell that synthesizes it. The actual synthetic product is an inactive precursor (zymogen) that can be activated later when and where needed (in the gut or to cause blood coagulation). Expert studied such enzyme precursors and compared the biological advantages of this means of control with the more usual ones of phosphorylation or allostery.
By 1990 it was becoming apparent that mass spectrometry might become a major tool of the protein chemist. Expert’s laboratory examined the power of electrospray mass spectrometry with proteins and peptides, then the relative merits of MALDI-TOF mass spectrometry. By 1995 it was clear that these tools were the methods of choice for the new field of "proteomics", whereby traces of an unknown protein could be identified. Convinced that every eukaryotic protein was posttranslationally modified, his lab focused on the use of mass spectrometry to identify a variety of posttranslational modifications, e.g., phosphorylation, acetylation, myristylation, etc.
Beginning with pancreatic trypsin and its precursor form trypsinogen, Expert’s studies ranged among its homologs (e.g. chymotrypsin) and blood coagulation factors (e.g. thrombin). In addition his studies probed carboxypeptidases and other proteases. Questions were raised regarding control, mechanism of action, homology and physiological role.
The central theme of Expert's studies has focused on the structure, function, control, and homology of proteins. In order to do so these proteins had to be purified. Hence many strategies were applied to separate proteins (and proteolytic fragments thereof). Progress was often limited by the need for better chromatographic methods.
Expert was a consultant for Amgen regarding protein chemistry research from 1985-1987.He was a consultant for Zymogenetics regarding protein chemistry from 1988-2000. Expert consulted for Bainbridge Sciences regarding protein chemistry from 1994-1996.
Expert may consult nationally and internationally, and is also local to the following cities: Seattle, Washington - Tacoma, Washington - Bellevue, Washington - Everett, Washington - Federal Way, Washington - Kent, Washington - Bellingham, Washington - Lakewood, Washington - Renton, Washington
|Year: 1959||Degree: Ph.D.||Subject: Enzymology/Biochemistry||Institution: University of Toronto|
|Year: 1953||Degree: M.S.||Subject: Biochemistry (focus on vitamins)||Institution: Purdue University|
|Year: 1951||Degree: B.Sc.||Subject: Food Chemistry||Institution: McGill University|
|Years: 2000 to Present||Employer: Undisclosed||Title: Professor Emeritus||Department: Biochemistry||Responsibilities: Expert retired as Departmental Chair in 2000 after 38 years of research and teaching.|
|Years: 1990 to 2000||Employer: University of Washington||Title: Acting Chair & Chair||Department: Biochemistry||Responsibilities: He gave leadership to a department of 20 faculty and about 150 individuals involved in biochemical research.|
|Years: 1962 to 2000||Employer: University of Washington||Title: Assistant Professor to Professor||Department: Biochemistry||Responsibilities: Expert taught biochemistry and directed research in protein chemistry.|
|Years: 1959 to 1962||Employer: University of Washington||Title: Postdoc||Department: Biochemistry||Responsibilities: He did research on protein chemistry and proteases.|
|Years: 1978 to 1982||Agency: NIH: Pysiol. Chem. Study Section||Role: Member; Chair 1980-82||Description: Expert reviewed grant applications in the area of physiological chemistry.|
|Years: 1969 to 2000||Agency: NIH||Role: Reviewer||Description: He reviewed numerous project sites for the NIH and made recommendations regarding merit.|
|Associations / Societies|
|Expert is a member of AAAS and ASBMB. He is a past member of the Protein Society and ABRF.|
|He was the first elected Secretary/Treasurer of the Protein Society (1987-90), and organized three national meetings. He was Chairman of the Department of Biochemistry at the University of Washington Medical School 1990-2000.|
|Awards / Recognition|
|Expert held the endowed Chair at the University of Washington from 1993-2000. He won the "Outstanding Teacher" award in 1997 and 2000, and the 2002 Edman Award from the MPSA.|
|Publications and Patents Summary|
|He has 237 peer-reviewed publications, largely on protein structure, function, and homologies. The most recent publications focused on the application of mass spectrometric methods to protein chemistry.|
|Expert Witness Experience|
|Expert was an expert witness in three cases concerning questions of patent infringement. Two involved disputes regarding the validity of amino acid sequence analysis data, one involved questions of protease specificity. He gave depositions twice. He testified as an expert witness in 2002 at a trial in a biotechnology patent dispute.|
|Training / Seminars|
|He has given perhaps 100 seminars on his research over the years at universities and institutes around the world. He taught protein chemistry in courses in Greece and Czechoslovakia. He taught graduate courses in protein chemistry and mass spectrometry at the University of Washington, and taught undergraduate biochemistry there for 40 years.|
|Other Relevant Experience|
| Expert served on various National Advisory Committees (e.g., the UCSF Mass Spectrometry Facility, NBRF Protein ID Resource, and Pacific Northwest National Lab).
He organized three national conferences (1000-1500 attendees) focused on proteins, and organized the International Conference on Methods in Protein Sequence Analysis in 1986.