Sequence differences between human muscle and liver cDNAs for UDPglucose pyrophosphorylase and kinetic properties of the recombinant enzymes expressed in Escherichia coli

Ronald G. Duggleby*, Yu Chyi Chao, Joyce G. Huang, Hwei-Ling Peng, Hwan You Chang

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

28 Scopus citations

Abstract

UDP-Glc pyrophosphorylase (EC 2.7.7.9) catalyses the interconversion of MgUTP plus Glc1P and UDP-Glc plus MgPP(i). Complementation of an Escherichia coli strain lacking this activity has allowed isolation of cDNA encoding this enzyme from a human muscle library. Two forms were identified and the nucleotide sequence of each was determined; they were found to differ only in the 5' region and we suggest that these arise from the use of a different first exon in the two transcripts. These nucleotide sequences are different from that of the cDNA which was isolated previously from a human liver library [Peng, H.-L. and Chang, H.-Y. (1993) FEBS Lett. 329, 153-158] and it is proposed that these liver and muscle forms are derived from different genes. The cDNA for muscle form I, muscle form II, the liver form, and the liver form fused to part of the lacZ gene were expressed in Escherichia coli and the kinetic properties of each enzyme were characterised. Muscle form I and the LacZ/liver fusion enzyme exhibit Michaelis-Menten kinetics towards all substrates while muscle form II has a sigmoidal dependence of rate upon the concentration of MgPP(i). The liver form shows Michaelis-Menten kinetics towards MgUTP. For the remaining three substrates, complex kinetics were observed involving a combination of sigmoidicity at low substrate concentration and partial inhibition at high substrate concentration.

Original languageEnglish
Pages (from-to)173-179
Number of pages7
JournalEuropean Journal of Biochemistry
Volume235
Issue number1-2
DOIs
StatePublished - 1 Jan 1996

Keywords

  • cDNA sequence
  • enzyme kinetics
  • galU
  • human isoenzymes
  • UDPglucose pyrophosphorylase

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