HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Meagher, J. L. Articles by Gettins, P. G. W. Search for Related Content PubMed PubMed Citation Articles by Meagher, J. L. Articles by Gettins, P. G. W. Social Bookmarking                      What's this?

J Biol Chem, Vol. 273, Issue 36, 23283-23289, September 4, 1998

Deconvolution of the Fluorescence Emission Spectrum of Human Antithrombin and Identification of the Tryptophan Residues That Are Responsive to Heparin Binding

Jennifer L. Meagher, Joseph M. Beechem^§, Steven T. Olson^¶, and Peter G. W. Gettins^¶

From the  Department of Biochemistry and Molecular Biology, College of Medicine and the ^¶ Center for Molecular Biology of Oral Diseases, College of Dentistry, University of Illinois at Chicago, Chicago, Illinois 60612 and the ^§ Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee, 37232

Heparin causes an allosterically transmitted conformational change in the reactive center loop of antithrombin and a 40% enhancement of tryptophan fluorescence. We have expressed four human antithrombins containing single Trp  Phe mutations and determined that the fluorescence of antithrombin is a linear combination of the four tryptophans. The contributions to the spectrum of native antithrombin at 340 nm were 8% for Trp-49, 10% for Trp-189, 19% for Trp-225, and 63% for Trp-307. Trp-225 and Trp-307 accounted for the majority of the heparin-induced fluorescence enhancement, contributing 37 and 36%, respectively. Trp-49 and Trp-225 underwent spectral shifts of 15 nm to blue and 5 nm to red, respectively, in the antithrombin-heparin complex. The blue shift for Trp-49 is consistent with partial burial by contact with heparin, whereas the red shift for Trp-225 and large enhancement probably result from increased solvent access upon heparin-induced displacement of the contact residue Ser-380. The enhancement for Trp-307 may result from the heparin-induced movement of helix H seen in the crystal structure. The time-resolved fluorescence properties of individual tryptophans of wild-type antithrombin were also determined using the four variants and showed that Trp-225 and Trp-307 experienced the largest change in lifetime upon heparin binding, providing support for the steady-state fluorescence deconvolution.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				J. Langdown, D. J. D. Johnson, T. P. Baglin, and J. A. Huntington Allosteric Activation of Antithrombin Critically Depends upon Hinge Region Extension J. Biol. Chem., November 5, 2004; 279(45): 47288 - 47297. [Abstract] [Full Text] [PDF]

				F. R. Melo, M. S. Pereira, D. Foguel, and P. A. S. Mourao Antithrombin-mediated Anticoagulant Activity of Sulfated Polysaccharides: DIFFERENT MECHANISMS FOR HEPARIN AND SULFATED GALACTANS J. Biol. Chem., May 14, 2004; 279(20): 20824 - 20835. [Abstract] [Full Text] [PDF]

				M. A. Jairajpuri, A. Lu, U. Desai, S. T. Olson, I. Bjork, and S. C. Bock Antithrombin III Phenylalanines 122 and 121 Contribute to Its High Affinity for Heparin and Its Conformational Activation J. Biol. Chem., April 25, 2003; 278(18): 15941 - 15950. [Abstract] [Full Text] [PDF]

				K. J. Belzar, A. Zhou, R. W. Carrell, P. G. W. Gettins, and J. A. Huntington Helix D Elongation and Allosteric Activation of Antithrombin J. Biol. Chem., March 1, 2002; 277(10): 8551 - 8558. [Abstract] [Full Text] [PDF]

				Z. Shriver, M. Sundaram, G. Venkataraman, J. Fareed, R. Linhardt, K. Biemann, and R. Sasisekharan Cleavage of the antithrombin III binding site in heparin by heparinases and its implication in the generation of low molecular weight heparin PNAS, September 12, 2000; 97(19): 10365 - 10370. [Abstract] [Full Text] [PDF]

				A. Futamura and P. G. W. Gettins Serine 380 (P14) right-arrow Glutamate Mutation Activates Antithrombin as an Inhibitor of Factor Xa J. Biol. Chem., February 11, 2000; 275(6): 4092 - 4098. [Abstract] [Full Text] [PDF]

				J. L. Meagher, S. T. Olson, and P. G. W. Gettins Critical Role of the Linker Region between Helix D and Strand 2A in Heparin Activation of Antithrombin J. Biol. Chem., January 28, 2000; 275(4): 2698 - 2704. [Abstract] [Full Text] [PDF]

				S. M. Van Patten, E. Hanson, R. Bernasconi, K. Zhang, P. Manavalan, E. S. Cole, J. M. McPherson, and T. Edmunds Oxidation of Methionine Residues in Antithrombin. EFFECTS ON BIOLOGICAL ACTIVITY AND HEPARIN BINDING J. Biol. Chem., April 9, 1999; 274(15): 10268 - 10276. [Abstract] [Full Text] [PDF]

				R. Bell, W. K. Stevens, Z. Jia, J. Samis, H. C. F. Cote, R. T. A. MacGillivray, and M. E. Nesheim Fluorescence Properties and Functional Roles of Tryptophan Residues 60d, 96, 148, 207, and 215 of Thrombin J. Biol. Chem., September 15, 2000; 275(38): 29513 - 29520. [Abstract] [Full Text] [PDF]

				O. Tcherkasskaya, V. E. Bychkova, V. N. Uversky, and A. M. Gronenborn Multisite Fluorescence in Proteins with Multiple Tryptophan Residues. APOMYOGLOBIN NATURAL VARIANTS AND SITE-DIRECTED MUTANTS J. Biol. Chem., November 10, 2000; 275(46): 36285 - 36294. [Abstract] [Full Text] [PDF]

				M. P. McGee and J. Liang Regulation of Glycosaminoglycan Function by Osmotic Potentials. MEASUREMENT OF WATER TRANSFER DURING ANTITHROMBIN ACTIVATION BY HEPARIN J. Biol. Chem., December 21, 2001; 276(52): 49275 - 49282. [Abstract] [Full Text] [PDF]