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Manuel E. Than, Gleb P. Bourenkov, Stefan Henrich, Karlheinz Mann, Wolfram Bode
The NC1 dimer of human placental basement membrane collagen IV: does a covalent crosslink exist?
Keywords: collagen IV, crosslink, Crystal structure, Edman analysis, NC1 domain
Triple-helical collagen IV protomers associate through their N- and C-termini, forming a three-dimensional network that provides basement membranes with mechanical strength. Within this network, the C-terminal non-collagenous (NC1) domains form tight dimeric junctions. Crystallographic analyses of isolated NC1 domains show two trimeric cap-like structures interacting via a large interface. Previously, for NC1 from human placenta type-IV collagen we described covalent ?1-?1 and ?2-?2 crosslinks between Met93 and Lys211 of opposing ?1(IV) and ?2(IV) NC1-chains, which further stabilize this interface and explain the occurrence of reduction-insensitive NC1-chain dimers. However, their existence was recently questioned, and we therefore analyzed NC1-domain dimers in more detail by biochemical and protein crystallographic methods. Short-exposure diffraction data show a clear electron density cross-connecting the respective residues, which gradually disappears with prolonged crystal irradiation. Sequence analyses of isolated tryptic peptides derived from denatured NC1 monomers and dimers indicate that only the dimers, but not the monomers, yield these chemically labile cross-linked peptides. These data clearly demonstrate the presence of reduction-resistant, but chemically and radiation-sensitive covalent crosslinks between the side chains of Met93 and Lys211 in human placenta type-IV collagen.
Biological Chemistry, Walter de Gruyter
Print ISSN: 1431-6730
Volume: 386, 08/2005
Pages: 759 - 766