No abstract

No abstract

An immunoglobulin light chain dimer with a large generic binding cavity was used as a host molecule for designing a series of peptide guest ligands. In a screening procedure peptides coupled to solid supports were systematically tested for binding activity by enzyme linked immunosorbent assays (ELISA). Key members of the binding series were synthesized in milligram quantities and diffused into crystals of the host molecule for X-ray analyses. These peptides were incrementally increased in size and affinity until they nearly filled the cavity. Progressive changes in binding patterns were mapped by comparisons of crystallographically refined structures of 14 peptide-protein complexes at 2.7 A resolution. These comparisons led to guidelines for ligand design and also suggested ways to modify previously established binding patterns. By manipulating equilibria involving histidine, for example, it was possible to abolish one important intramolecular interaction of the bound ligand and substitute another. These events triggered a change in conformation of the ligand from a compact to an extended form and a comprehensive change in the mode of binding to the protein. In dipeptides of histidine and proline, protonation of both imidazolium nitrogen atoms was used to program an end-to-end reversal of the direction in which the ligand was inserted into the binding cavity. Peptides cocrystallized with proteins produced complexes somewhat different in structure from those in which ligands were diffused into preexisting crystals. In such a large and malleable cavity, space utilization was thus different when a ligand was introduced before the imposition of crystal packing restraints.

Multipin peptide synthesis has been employed to produce biotinylated 11-mer phosphopeptides that account for every tyrosine residue in insulin receptor substrate-1 (IRS-1) and the cytoplasmic domains of the insulin-, epidermal growth factor-, platelet-derived growth factor-and basic fibroblast growth factor receptors. These phosphopeptides have been screened for their capacity to bind to the SH2 domains of Shc and Grb

Virus-specific or group-specific antibody probes to potyviruses can be produced by targeting the immune response to the virus-specific, N-terminal region of the capsid protein (29-95 amino acids depending on the virus) or to the conserved core region (216 amino acids) of the capsid protein, respectively. Immunochemical analysis of overlapping, synthetic octapeptides covering the capsid protein of the Johnsongrass strain of Johnsongrass mosaic virus (JGMV-JG) has delineated the peptide sequences recognized by five polyclonal rabbit antisera and two mouse monoclonal antibodies (mAbs). The antibodies characterized were (i) three virus-specific rabbit polyclonal antisera and one virus-specific mouse mAb (1/25) raised against native virus particles, (ii) one polyclonal antiserum raised against trypsin-derived core particles of JGMV-JG, (iii) one group-specific polyclonal antiserum raised against the denatured, truncated coat protein from trypsinderived core particles of JGMV-JG, and (iv) one group-specific mouse mAb (1/16) raised against native virus particles. The two epitopes seen by mAb 1/25 occurred at residues 18-27 and 43-52 and overlapped with the two major epitopes seen by the virus-specific polyclonal antiserum. The group-specifilc epitope seen in JGMV-JG by mAb 1/16 was also recognized strongly in potato virus Y. the type member of the potyvirus group. The multiple epitopes seen by the cross-reactive polyclonal antisera were distributed across the entire core region of the coat protein and their relative antibody binding responses varied between JGMV-JG, potato virus Y, and six other distinct potyviruses.Plant diseases are estimated to be responsible for economic losses worldwide of $60 billion per annum (1). The most important pathogens are fungi, with plant viruses the second most important group of infectious agents. Of the 28 plant virus groups or families the potyvirus group is the largest. It contains at least 175 independent members and accounts for about 30% of all viruses known to infect plant species around the world (2, 3). Potyvirus particles are flexuous rods, 729-900 nm long and -11 nm in diameter, and consist of up to 2000 subunits of a single protein species (4).Successful control and eradication of plant virus infections is dependent on the availability of simple, reliable methods for plant virus detection and identification. To date, this has been difficult to achieve for potyviruses due to the large size of the group, the vast variation between members, and the lack of satisfactory taxonomic parameters to distinguish independent viruses from related strains (2-4). Thus, there is a real need to evaluate new criteria for the identification and classification of potyviruses.During investigations on the structural characterization of the coat proteins of potyviruses we made the following observations that have implications for potyvirus detection and classification: (i) distinct potyviruses exhibit coat protein sequence homology of 38-71% with major differences in the length (29-95 re...