p-chloromercuribenzoate-induced inactivation and partial unfolding of porcine heart lactate dehydrogenase

Y. B. Zheng, B. Y. Chen, X. C. Wang

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

Purified porcine heart lactate dehydrogenase was inactivated and partially unfolded with p-chloromercuribenzoate (pCMB). With the increase of pCMB/enzyme ratio the enzyme was gradually inhibited till almost completely inactivated at the pCMB/enzyme ratio of 20: 1. Native polyacrylamide gel electrophoresis showed that with the increase of pCMB/enzyme ratio the bands of native enzyme decreased till completely vanished. Meanwhile inactive multiple bands emerged and became thicker, which implied that lactate dehydrogenase became loose. The conformational changes of the enzyme molecule modified with pCMB were followed using fluorescence emission, ultraviolet difference, and circular dichroism (CD) spectra. Increasing pCMB concentration resulted in the decrease of fluorescence emission intensity. The ultraviolet difference spectra of the enzyme modified with pCMB exhibited an increasing absorbance in the vicinity of 240 nm with the increasing concentration of the inhibitor. The changes of the fluorescence and ultraviolet difference spectra reflected the conformational changes of the enzyme. The CD spectrum changes of the enzyme showed that its secondary structure changed as well. These results suggest that pCMB not only inhibits this enzyme but also influences its conformation (partial unfolding).

Original languageEnglish (US)
Pages (from-to)583-587
Number of pages5
JournalBiochemistry (Moscow)
Volume67
Issue number5
DOIs
StatePublished - May 2002
Externally publishedYes

Keywords

  • Inactivation
  • Lactate dehydrogenase
  • Unfolding
  • p-chloromercuribenzoate

ASJC Scopus subject areas

  • Biochemistry

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