Determination of Sulfated Glycosaminoglycan Binding Sites Within Collagen Type XI Using Surface Plasmon Resonance and Nuclear Magnetic Resonance Spectroscopy
Dr. Julia Oxford and Owen McDougal
Type XI Collagen is a minor constituent of the extracellular matrix of cartilage and is essential in the regulation of collagen fibril assembly and diameter. The alpha 1 chain of Collagen XI (Collagen a1(XI) ) contains a variable region that is modulated by alternative splicing in a tissue-specific and developmental manner. Preliminary data suggests that some Collagen XI isoforms interact with sulfated gylcosaminoglycans. Sulfated glycosaminoglycans are unbranched polysaccharides containing a disaccharide subunit. Chondroitin sulfate is a sulfated glycosaminoglycan which contains repeating disaccharide subunits composed of D-glucuronate and N-acetyl-D-glalactosamine sulfate. Chondroitin sulfate is located primarily in the extracellular matrix of cells. The function of chondroitin sulfate is not well understood, but has been shown to interact with proteins, and may play structural roles within the body. Biophysical methods will be used to assess the binding interactions between Collagen a1(XI) and the glycosaminoglycan chondroitin sulfate. Collagen a1(XI) isoform V1B was expressed in Escherichia coli and purified using affinity chromatography. Peptides pertaining to potential glycosaminoglycan binding sites in the variable region of the V1B isoform of Collagen a1(XI) were synthesized. Surface plasmon resonance spectroscopy was used to determine the binding interactions between Collagen a1(XI) peptides and chondroitin sulfate glycosaminoglycans. Nuclear magnetic resonance (NMR) spectroscopy was used to determine the structure of the V1B peptides and the physical interactions occuring between the peptides and chondroitin sulfate.