Rat lysyl hydroxylase: molecular cloning, mRNA distribution and expression in a baculovirus system

Abstract

A cDNA library from rat lung was screened with a chicken lysyl hydroxylase cDNA, and several overlapping rat lysyl hydroxylase cDNAs were isolated. The complete cDNA was 91 and 77% identical, respectively, to the human and chicken lysyl hydroxylase cDNAs at the protein level. By Northern blot, the rat lysyl hydroxylase cDNA recognized a single 3.2 kb mRNA that was present in a wide variety of rat tissues. In order to further confirm the identity of this cDNA, the cDNA was expressed in insect cells via a baculovirus vector. These cells produced an 85 kDa protein with lysyl hydroxylase activity. The recombinant lysyl hydroxylase had a specific activity and Km values for its substrates that were similar to those of the enzyme isolated from chick embryos. The fact that this single lysyl hydroxylase cDNA encodes a protein sufficient for lysyl hydroxylase activity is consistent with previous biochemical findings that lysyl hydroxylase only requires a single type of subunit for its activity.