Characterisation of lysyl hydroxylases and their role in collagen cross-linking

Defects in collagen synthesis and metabolism can lead to a number of clinically significant diseases, such as osteoarthritis, osteoporosis, osteogenesis imperfecta, and diseases associated with tissue fibrosis, such as cardiovascular disease and liver cirrhosis. Fibrillogenesis, the formation of collagen fibrils, is a complex process requiring extensive post-translational modifications of procollagen to form mature collagen fibrils. Crucial among them is the hydroxylation of lysine residues in both the telopeptide and triple helical regions of the procollagen polypeptide that directs intra- and inter-molecular collagen cross-linking. Hydroxylation within the helical domain is catalysed by the enzyme lysyl hydroxylase (LH) (procollagen-lysine 2-oxoglutarate, 5-dioxygenase; PLOD) whereas telopeptide lysyl hydroxylation was suggested to be accomplished by a novel enzyme. Originally, LH was believed to be a single protein, but three isoforms (LH1, LH2 and LH3) have been characterised in human, mouse, including two splice variant of LH2 (LH2b (long) and LH2a (short)). Only one isoform had been identified in the rat. This thesis focuses on the role of lysyl hydroxylases in hydroxylation of lysine residues in collagen by examination of their molecular and biochemical characteristics, in relation to health and disease. Evidence that one of the LH, rather than a novel enzyme, is telopeptide lysyl hydroxylase (TLH) is presented. Vitamin C, an important vitamin only available from the diet, plays an important role in collagen biosynthesis and pyridinium cross-link production. Vitamin C supplementation increased collagen production and cross-link production in a cell culture model system, and altered the expression of LH gene expression. The effects of a number of inhibitors of collagen synthesis were also examined for their effect on LH gene expression, and in particular TLH.EThOS - Electronic Theses Online ServiceGBUnited Kingdo