Evidence for the presence of a serine proteinase(s) associated with the microsomal membranes of rat liver

Kazuhiro Sogawa, Kenji Takahashi

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Neutral proteinase activity in subcellular fractions of rat liver was measured with heat-denatured casein as a substrate before and after repeated freezing and thawing, followed by extraction of each fraction with 1 M KC1. By this treatment, most of the proteinase activity in each fraction was solubilized except for that in the microsomal fraction. More than 50% of the neutral proteinase activity in the microsomal fraction remained insoluble, which indicated that the 1 M KCI-insoluble neutral proteinase activity was predominantly localized in the microsomal fraction. The activity was present in both the rough and smooth microsomes.The neutral proteinase(s) was largely solubilized from the 1 M KCI-washed microsomal fraction by 0.5 M NaCl containing 0.5 % deoxycholate and 0.25 % cholate. The solubilized pro-teinase(s) was eluted from a Sepharose CL-6B column as a single but rather broad peak, giving a molecular weight of about 84,000. It showed maximal activity at pH 8.0 toward heat-denatured casein as a substrate. It was markedly inhibited by diisopropyl phosphorofluoridate, but was not significantly inhibited by chymostatin, soybean trypsin inhibitor, p-chloromer-curibenzoate or EDTA. o-Phenanthroline was somewhat inhibitory. Among urea-denatured proteins tested as substrates, calf thymus histone was hydrolyzed most rapidly, followed by casein and hemoglobin, whereas ovalbumin, bovine serum albumin, and γ-globulin were not hydrolyzed to any significant extent.These results demonstrate that rat liver microsomes contain a unique serine proteinase(s) firmly bound to their membranes.

Original languageEnglish
Pages (from-to)763-770
Number of pages8
JournalJournal of biochemistry
Issue number4
Publication statusPublished - 1978 Oct
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology


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