Rat lysyl hydroxylase

molecular cloning, mRNA distribution and expression in a baculovirus system

Lucas C. Armstrong, Jerold A Last

Research output: Contribution to journalArticle

27 Citations (Scopus)

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.

Original languageEnglish (US)
Pages (from-to)93-102
Number of pages10
JournalBBA - Gene Structure and Expression
Volume1264
Issue number1
DOIs
StatePublished - Oct 17 1995

Fingerprint

2-Oxoglutarate 5-Dioxygenase Procollagen-Lysine
Baculoviridae
Cloning
Molecular Cloning
Rats
Complementary DNA
Messenger RNA
Chickens
Proteins
Chick Embryo
Gene Library
Northern Blotting
Insects

Keywords

  • (Rat)
  • Baculovirus vector
  • cDNA
  • Lysyl hydroxylase
  • Molecular cloning
  • mRNA distribution

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Genetics
  • Molecular Biology
  • Structural Biology

Cite this

Rat lysyl hydroxylase : molecular cloning, mRNA distribution and expression in a baculovirus system. / Armstrong, Lucas C.; Last, Jerold A.

In: BBA - Gene Structure and Expression, Vol. 1264, No. 1, 17.10.1995, p. 93-102.

Research output: Contribution to journalArticle

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AB - 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.

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