Photochemical crosslinking of bacteriophage T4 single‐stranded DNA‐binding protein (gp32) to oligo‐p(dT)8: Identification of phenylalanine‐183 as the site of crosslinking
- 1 January 1988
- journal article
- research article
- Published by Wiley in Proteins-Structure Function and Bioinformatics
- Vol. 4 (1) , 1-6
- https://doi.org/10.1002/prot.340040103
Abstract
Using ultraviolet light, both the 33,000-dalton single-standard DNA-binding protein from T4 bacteriophage (gp32)as well as a 25,000-dalton limited trypsin cleavage product of gp32 (core gp32*) that retains high affinity for single-stranded DNA can be crosslinked to an oligodeoxynucleotide, p(dT)8. After photolysis, a single tryptic peptide crosslinked to p(dT)8 was isolated by anion-exchange high-performance liquid chromatography. Gas-phase sequencing of this modified peptide gave the following sequence: Gln-Val-Ser-Gly-(X)-Ser-Asn-Tyr-Asp-Glu-Ser-Lys, which corresponds to residues 179–190 in gp32. Based on the absence of the expected phenylthiohydantoin derivative of phenylalanine 183 at cycle 5 (X) we infer that crosslinking has occurred at this position and that phenylalanine 183 is at the interface of the gp32:P(dT)8 complex in an orientation that allows covalent bond formation with the thymine radical produced by ultraviolet irradiation.Keywords
This publication has 19 references indexed in Scilit:
- The function of zinc in gene 32 protein from T4Biochemistry, 1987
- High-performance liquid chromatographic peptide mapping and amino acid analysis in the sub-nanomole rangeJournal of Chromatography A, 1986
- Cloning of T4 gene 32 and expression of the wild-type protein under lambda promoter PL regulation in Escherichia coli.Proceedings of the National Academy of Sciences, 1986
- Proton NMR (500 MHz) identification of aromatic residues of gene 32 protein involved in DNA binding by use of protein containing perdeuterated aromatic residues and by site-directed mutagenesisBiochemistry, 1986
- SINGLE-STRANDED DNA BINDING PROTEINS REQUIRED FOR DNA REPLICATIONAnnual Review of Biochemistry, 1986
- Maximizing gene expression from plasmid vectors containing the lambda PL promoter: strategies for overproducing transcription termination factor rho.Proceedings of the National Academy of Sciences, 1985
- Proton NMR (500 MHz) of gene 32 protein-oligonucleotide complexesBiochemistry, 1984
- Refined structure of the gene 5 DNA binding protein from bacteriophage fdJournal of Molecular Biology, 1983
- Interactions of bacteriophage T4-coded gene 32 protein with nucleic acidsJournal of Molecular Biology, 1981
- Physicochemical properties of DNA binding proteins. Gene 32 protein of T4 and Escherichia coli unwinding proteinBiochemistry, 1975