Characterization of the effects of the disease causing mutations on the triple helix conformation using recombinant collagen fragments from E coli.

Collagens are key factors for tissue integrity and function, yet the molecular interactions involving collagen remain poorly understood. All collagens share the same triple helix structure consisting of three polypeptide chains with the distinct Gly ‐X‐Y repeating sequence pattern (where X and Y can be any amino acids). The triple helices further assemble into different supramolecular structures which confer the diverse biological functions of collagen. Characterization of molecular interactions of collagen has been difficult because of its large sizes, often containing more than 1000 amino acids per chain. Consequently, many of the presently available research tools are difficult and cumbersome to use. Short synthetic peptides used to study the conformation of triple helix normally do not exhibit the ability to further associate into higher order assemblies, and are generally considered poor models for collagens in tissues. In this presentation, we will report the progress of our approach to this challenge using recombinant collagen fragments from E coli with defined sizes We have created an E coli clone to produce cross linked triple helix fragments modeling the region of 877–893 of the α1 chain of type I collagen. Two disease causing mutations in this region are being introduced by mutagenesis. Biophysical characterizations of these fragments are focused on demonstrating the correlations of the regional conformational flexibility of the triple helix with the severity of the disease caused by different Gly substituting mutations. This work is supported in part by NSF grant DBI0521709, NIH grant 3S06‐GM060654‐06S1, PSC‐CUNY grant 0000, , and the Gender Equity program at Hunter (NSF grant 0000).