Biophysical Journal 69: 20-29 (1995)
© 1995 the Biophysical Society

This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Hasselbacher, C A Articles by Polikarpov, I Search for Related Content PubMed PubMed Citation Articles by Hasselbacher, C A Articles by Polikarpov, I

Environments of the four tryptophans in the extracellular domain of human tissue factor: comparison of results from absorption and fluorescence difference spectra of tryptophan replacement mutants with the crystal structure of the wild-type protein.

Department of Biochemistry, Mount Sinai School of Medicine, New York, New York 10029, USA.

ABSTRACT

The local environments of the four tryptophan residues of the extracellular domain of human tissue factor (sTF) were assessed from difference absorption and fluorescence spectra. The difference spectra were derived by subtracting spectra from single Trp-to-Phe or Trp-to-Tyr replacement mutants from the corresponding spectrum of the wild-type protein. Each of the mutants was capable of enhancing the proteolytic activity of factor VIIa showing that the mutations did not introduce major structural changes, although the mutants were more susceptible to denaturation by guanidinium chloride. The difference spectra indicate that the Trp residues are buried to different extents within the protein matrix. This evaluation was compared with the x-ray crystal structure of sTF. There is excellent agreement between predictions from the difference spectra and the environments of the Trp residues observed in the x-ray crystal structure, demonstrating that difference absorption and particularly fluorescence spectra derived from functional single-Trp replacement mutants can be used to obtain information about the local environments of individual Trp residues in multi-tryptophan proteins.

This article has been cited by other articles:

				J. L. Meagher, J. M. Beechem, S. T. Olson, and P. G. W. Gettins Deconvolution of the Fluorescence Emission Spectrum of Human Antithrombin and Identification of the Tryptophan Residues That Are Responsive to Heparin Binding J. Biol. Chem., September 4, 1998; 273(36): 23283 - 23289. [Abstract] [Full Text] [PDF]