Identifying the SPARC binding sites on collagen I and procollagen I by atomic force microscopy.
SPARC (secreted protein acidic and rich in cysteine) is a matricellular protein associated with the extracellular matrix (ECM). It has been found that the production of collagen I is a requisite for the association of SPARC with ECM, and studies with SPARC-null mice indicate that SPARC plays a role in modifying the structure of collagen fibers. It is not known, however, whether SPARC interacts with the collagen I precursor, procollagen I. In this study, the binding of SPARC to collagen I and procollagen I was verified by surface plasmon resonance. The SPARC-binding sites on collagen I and procollagen I were identified by directly visualizing their complexes using tapping-mode atomic force microscopy (TM-AFM). The characteristic chain end feature in collagen I is not readily detected by AFM, so unambiguous location of the binding sites relative to the C- or N-termini is difficult. In contrast, procollagen I, with its large globular C-propeptide, permits easy identification of the C-terminus. Histograms were constructed and compared based on the distances of the bound SPARC to the C-terminus of procollagen I and to the closest end of collagen I. There is a broad distribution of SPARC binding sites on procollagen I with the most preferred binding region located approximately 1/3 from the C-terminus. Characterization of the SPARC-binding sites on collagen I and procollagen I provides useful information for further understanding of the functional implications of their interactions.
Wang
Hua
H
Department of Chemical Engineering, University of Washington, Seattle, Washington 98195, USA.
Fertala
Andrzej
A
Ratner
Buddy D
BD
Sage
E Helene
EH
Jiang
Shaoyi
S
D000818:Animals; D001665:Binding Sites; D024042:Collagen Type I / Q000737:chemistry / Q000378:metabolism*; D018625:Microscopy, Atomic Force / Q000379:methods*; D015676:Osteonectin / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats
D024042:Collagen Type I; D015676:Osteonectin