CORE--PEPTIDE SYNTHESIS AND CONFORMATION FACILITY

Wistar Institute, Philadelphia PA

Investigators

Linked publications & trials

Abstract

Description: The Peptide Synthesis and Conformation Analysis Facility serves the Wistar Cancer Center investigators with the design, synthesis, and secondary structural analysis of peptides. Unmodified peptides as well as several types of peptide analogues are prepared in the facility by solid-phase chemical synthesis methods. A special emphasis of the facility is production, purification, and characterization of modified synthetic peptides, including phosphopeptides, glycopeptides, biotinylated peptides, fatty acid-coupled peptides, isomerized peptides, and those peptide analogues for which the appropriately protected monomers (amino acid residues) are either commercially available or can be prepared by the facility. The expertise of the facility in complex organic chemistry allows it to provide unique modified synthetic peptides, which are analyzed by chromatography, mass spectroscopy, and amino acid analysis to ensure accurate synthesis. Upon request, the peptides are purified by reversed-phase HPLC and coupled to carrier proteins that are used to enhance antigenicity. The conformation of peptides in solution is determined by circular dichroism (CD), Fourier-transform infrared spectroscopy (FTIR), and computer-aided molecular modeling. Since the last Wistar review, this facility has undergone major changes in personnel, instrumentation, and available technology. The facility is included in the Center because of the scientific interest in the use of peptides and a better understanding of their function(s). The long-term goal of the facility is to assist the Center investigators with all aspects of peptide use in their research. To this end, the facility also provides consultation concerning the selection, length, specific requirements, and conjugation strategies for synthesizing the peptide sequences. Since the last review, the facility has been transferred to the supervision of Dr. Laszlo Otvos, who joined the staff of Wistar in 1990 and is now a member of the Structural Biology Program. Dr. Otvos?s laboratory and the facility recently moved into a completely remodeled wing of the Institute that houses the Structural Biology Center. This new laboratory was specifically designed for organic chemistry, and especially for peptide chemistry. The Peptide Synthesis Facility occupies 1200 square feet of space that is integrated with Dr. Otvos' research laboratory. The Conformation Analysis Laboratory is located across the corridor. The facility is available to all Center investigators on a first-come, first served basis. The equipment dedicated to the Peptide Synthesis Facility includes a Rainin PS3 peptide synthesizer and Waters and Beckman gradient HPLC systems, lyophilizer, sonic dismembrator, and high speed mixer. The following equipment is dedicated to the Conformation Analysis Facility: Jasco J720 Circular Dichrograph, Nicolet Impact 400 Fourier-transform Infrared Spectrophotometer, and a Silicon Graphics 240 Workstation. An additional peptide synthesizer (MilliGen 9050), two HPLCs, a Silicon Graphics computer, dedicated for Dr. Otvos' research, are available to the facility when needed. The total number of Center users of the facility in the past three years is 18, 15 of whom have had peer-reviewed grant support. The number of peptides synthesized has gone from seven in 1993 to thirty three in 1995, although it is stated that currently two peptides per week are being synthesized. It should be noted that only about half of peptides synthesized are purified by the facility. During the 1993-95 period, a total of 72 projects involving synthesis of 213 peptides were supported by the facility. Of these, 25 projects involving 69 peptides were produced for Cancer Center investigators. In the past year, 25 percent of the usage has been for peer reviewed projects.