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

J. Biol. Chem., Vol. 283, Issue 48, 33310-33320, November 28, 2008

This Article Full Text Full Text (PDF) Supplemental Data All Versions of this Article: 283/48/33310    most recent M801287200v1 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 Becerra, S. P. Articles by Hollyfield, J. G. Search for Related Content PubMed PubMed Citation Articles by Becerra, S. P. Articles by Hollyfield, J. G. Social Bookmarking                      What's this?

Pigment Epithelium-derived Factor Binds to Hyaluronan

MAPPING OF A HYALURONAN BINDING SITE^*

S. Patricia Becerra¹, L. Alberto Perez-Mediavilla, John E. Weldon, Silvia Locatelli-Hoops, Preenie Senanayake^¶, Luigi Notari, Vicente Notario^||, and Joe G. Hollyfield^¶

From the NEI, National Institutes of Health, Bethesda, Maryland 20892, the Center for Applied Medical Research, University of Navarra, Pamplona, Spain, the ^¶Department of Ophthalmology, Cleveland Clinic Lerner College of Medicine, Cleveland, Ohio 44195, and the ^||Radiation Medicine Department, Georgetown University Medical Center, Washington, D. C. 20007

Pigment epithelium-derived factor (PEDF) is a multifunctional serpin with antitumorigenic, antimetastatic, and differentiating activities. PEDF is found within tissues rich in the glycosaminoglycan hyaluronan (HA), and its amino acid sequence contains putative HA-binding motifs. We show that PEDF coprecipitation with glycosaminoglycans in media conditioned by human retinoblastoma Y-79 cells decreased after pretreatments with hyaluronidase, implying an association between HA and PEDF. Direct binding of human recombinant PEDF to highly purified HA was demonstrated by coprecipitation in the presence of cetylpyridinium chloride. Binding of PEDF to HA was concentration-dependent and saturable. The PEDF-HA interactions were sensitive to increasing NaCl concentrations, indicating an ionic nature of these interactions and having affinity higher than PEDF-heparin. Competition assays showed that PEDF can bind heparin and HA simultaneously. PEDF chemically modified with fluorescein retained the capacity for interacting with HA but lacked heparin affinity, suggesting one or more distinct HA-binding regions on PEDF. The HA-binding region was examined by site-directed mutagenesis. Single-point and cumulative alterations at basic residues within the putative HA-binding motif K189A/K191A/R194A/K197A drastically reduced the HA-binding activity without affecting heparin- or collagen I binding of PEDF. Cumulative alterations at sites critical for heparin binding (K146A/K147A/R149A) decreased HA affinity but not collagen I binding. Thus these clusters of basic residues (BXBXXBXXB and BX₃AB₂XB motifs) in PEDF are functional regions for binding HA. In the spatial PEDF structure they are located in distinct areas away from the collagen-binding site. The HA-binding activity of PEDF may contribute to deposition in the extracellular matrix and to its reported antitumor/antimetastatic effects.

Received for publication, February 19, 2008 , and in revised form, September 15, 2008.

^* This work was supported, in whole or in part, by National Institutes of Health NEI Intramural Research Program. This work was also supported by the Educational and Cultural Department of the "Gobierno de Navarra," Spain (to A. P.-M.); by Grants EY14240-1 and EY 15638, Foundation for Fighting Blindness, and a Challenge Grant from Research to Prevent Blindness to the Department of Ophthalmology, Cleveland Clinic Lerner College of Medicine (to J. G. H.); and by Grant RO1 CA64472 (to V. N.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental text and Figs. S1–S5.

¹ To whom correspondence should be addressed: NEI, NIH, Bldg. 7, Rm. 304, 7 Memorial Drive, Bethesda, MD 20892-0706. Tel.: 301-496-6514; Fax: 301-451-5420; E-mail: becerrap{at}nei.nih.gov.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				A. Bernard, J. Gao-Li, C.-A. Franco, T. Bouceba, A. Huet, and Z. Li Laminin Receptor Involvement in the Anti-angiogenic Activity of Pigment Epithelium-derived Factor J. Biol. Chem., April 17, 2009; 284(16): 10480 - 10490. [Abstract] [Full Text] [PDF]