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Journal of Virology, July 2004, p. 6766-6774, Vol. 78, No. 13
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.13.6766-6774.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Identification of the Human Rhinovirus Serotype 1A Binding Site on the Murine Low-Density Lipoprotein Receptor by Using Human-Mouse Receptor Chimeras

Barbara Herdy, Luc Snyers, Manuela Reithmayer, Peter Hinterdorfer, and Dieter Blaas^*

Max F. Perutz Laboratories, University Departments at the Vienna Biocenter, Department of Medical Biochemistry, University of Vienna, A-1030 Vienna, Austria

Received 7 November 2003/ Accepted 22 March 2004

Human rhinovirus serotype 1A (HRV1A) binds more strongly to the mouse low-density lipoprotein receptor (LDLR) than to the human homologue (M. Reithmayer, A. Reischl, L. Snyers, and D. Blaas, J. Virol. 76:6957-6965, 2002). Here, we used this fact to determine the binding site of HRV1A by replacing selected ligand binding modules of the human receptor with the corresponding ligand binding modules of the mouse receptor. The chimeric proteins were expressed in mouse fibroblasts deficient in endogenous LDLR and LDLR-related protein, both used by minor group HRVs for cell entry. Binding was assessed by virus overlay blots, by immunofluorescence microscopy, and by measuring cell attachment of radiolabeled virus. Replacement of ligand binding repeat 5 of the human LDLR with the corresponding mouse sequence resulted in a substantial increase in HRV1A binding, whereas substitution of repeats 3 and 4 was without effect. Replacement of human receptor repeats 1 and 2 with the murine homologues also increased virus binding. Finally, murine receptor modules 1, 2, and 5 simultaneously introduced into the human receptor resulted in HRV1A binding indistinguishable from mouse wild-type receptor. Thus, repeats 1 and/or 2 and repeat 5 are involved in HRV1A attachment. Changing CDGGPD in the acidic cluster of module 5 in the human receptor to CDGEAD present in the mouse receptor led to substantially increased binding of HRV1A, indicating an important role of the glutamate residue in HRV1A recognition.

Present address: Institute of Histology and Embryology, University of Vienna, A-1090 Vienna, Austria.

Present address: Institute for Biophysics, J. Kepler University, A-4040 Linz, Austria.

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