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Journal of Virology, October 2003, p. 11072-11081, Vol. 77, No. 20
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.20.11072-11081.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Identification of a Heparin-Binding Motif on Adeno-Associated Virus Type 2 Capsids{dagger}

A. Kern,1 K. Schmidt,1 C. Leder,1 O. J. Müller,1 C. E. Wobus,1,{ddagger} K. Bettinger,2 C. W. Von der Lieth,2 J. A. King,1,§ and J. A. Kleinschmidt1*

Forschungsschwerpunkt Angewandte Tumorvirologie,1 Zentrale Spektroskopie, Deutsches Krebsforschungszentrum, 69120 Heidelberg, Germany2

Received 8 May 2003/ Accepted 15 July 2003

Infection of cells with adeno-associated virus (AAV) type 2 (AAV-2) is mediated by binding to heparan sulfate proteoglycan and can be competed by heparin. Mutational analysis of AAV-2 capsid proteins showed that a group of basic amino acids (arginines 484, 487, 585, and 588 and lysine 532) contribute to heparin and HeLa cell binding. These amino acids are positioned in three clusters at the threefold spike region of the AAV-2 capsid. According to the recently resolved atomic structure for AAV-2, arginines 484 and 487 and lysine 532 on one site and arginines 585 and 588 on the other site belong to different capsid protein subunits. These data suggest that the formation of the heparin-binding motifs depends on the correct assembly of VP trimers or even of capsids. In contrast, arginine 475, which also strongly reduces heparin binding as well as viral infectivity upon mutation to alanine, is located inside the capsid structure at the border of adjacent VP subunits and most likely influences heparin binding indirectly by disturbing correct subunit assembly. Computer simulation of heparin docking to the AAV-2 capsid suggests that heparin associates with the three basic clusters along a channel-like cavity flanked by the basic amino acids. With few exceptions, mutant infectivities correlated with their heparin- and cell-binding properties. The tissue distribution in mice of recombinant AAV-2 mutated in R484 and R585 indicated markedly reduced infection of the liver, compared to infection with wild-type recombinant AAV, but continued infection of the heart. These results suggest that although heparin binding influences the infectivity of AAV-2, it seems not to be necessary.

* Corresponding author. Mailing address: Angewandte Tumorvirologie, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 242, 69120 Heidelberg, Germany. Phone: 49 6221 424978. Fax: 49 6221 424962. E-mail: J.Kleinschmidt{at}dkfz.de.

{dagger} This article is dedicated to Harald zur Hausen on the occasion of his retirement as head of the German Cancer Research Center with gratitude and appreciation for 20 years of leadership.

{ddagger} Present address: Department of Pathology, School of Medicine, Washington University, St. Louis, MO 63122.

§ Present address: MRC Centre for Inflammation Research, Edinburgh University Medical School, Edinburgh EH8 9AG, Scotland.

Journal of Virology, October 2003, p. 11072-11081, Vol. 77, No. 20
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.20.11072-11081.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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