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Histidine-tryptophan interactions in T4 lysozyme: 'anomalous' pH dependence of fluorescence.

Van Gilst, M and Hudson, B S (1996) Histidine-tryptophan interactions in T4 lysozyme: 'anomalous' pH dependence of fluorescence. Biophysical chemistry, 63 (1). pp. 17-25. ISSN 0301-4622

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Article URL: http://dx.doi.org/10.1016/S0301-4622(96)02181-3

Abstract

A variant of T4 lysozyme which contains only a single tryptophan residue (at position 138) has been prepared (W126Y/W158Y designated 'YWY'). Two additional mutations to YWY have been prepared involving replacement of glutamine 105, which hydrogen bonds to the indole N-H of trp 138 in wild type, with either a histidine (YWY/Q105H) or an alanine (YWY/Q105A). The fluorescence properties of these two species are investigated as a function of pH. YWY/Q105A exhibits essentially a single exponential fluorescence decay (5% tau = 0.35 ns 95% tau = 5 ns) and almost no pH dependence in steady state or time resolved fluorescence behavior. In contrast, YWY/Q105H exhibits complex fluorescence decay over the entire pH range used in these experiments. As the pH is lowered from 8 to 4, there is an increase in the quantum yield and a change in the average lifetime (from 2.0 to 3.1 ns). Using this data, the pKa of histidine 105 has been determined to be 5.9. These results are contrasted to those from other proteins which show a pH dependent tryptophan fluorescence associated with a neighboring histidine or other residue. Quenching behavior in terms of the stereochemistry of the tryptophan-histidine interaction and implications of these results for current models of complex fluorescence behavior of single tryptophan proteins are also discussed.

Item Type: Article
DOI: 10.1016/S0301-4622(96)02181-3
PubMed ID: 8981748
Grant Numbers: GM36578
Keywords or MeSH Headings: Fluorescence; Histidine/metabolism; Hydrogen-Ion Concentration; Models, Molecular; Muramidase/metabolism; Mutagenesis/genetics; T-Phages/enzymology; Tryptophan/genetics/metabolism;
Subjects: Molecules > Proteins > Enzymes
Molecules > Proteins
Depositing User: Library Staff
Date Deposited: 24 Sep 2008 21:59
Last Modified: 07 May 2010 20:34
URI: http://authors.fhcrc.org/id/eprint/60

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