Purification and characterization of a Ca2+-activated thiol protease from Drosophila melanogaster

Marianna Pintér, P. Friedrich, Peter Friedrich

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

A Ca2+-activated thiol protease was purified from Drosophila melanogaster. The procedure involves Phenyl-Sepharose, Reactive Red-Agarose and Q-Sepharose fast flow (or MonoQ) chromatographic steps. The enzyme eluting from Q-Sepharose fast flow seems to be homogeneous as judged by silver staining on SDS-PAGE: it consists of a single polypeptide chain of Mr,app = 94K and pI = 5.46. The proteolytic activity of the purified enzyme is absolutely Ca2+-dependent, characterized by 0.6 mM free Ca2+ at half-maximal activity. Ca2+ ions cannot be replaced effectively by the divalent cations Mg2+, Mn2+, Zn2+, Ba2+, and Cd2+. The enzyme shows the inhibitor pattern of thiol proteases. Human recombinant calpastatin (domain I) completely inhibits the enzyme at a nearly 1:1 molar ratio. Several of these properties resemble those of vertebrate calpain II. However, various attempts to detect a small subunit of Mr ∼30K, common with vertebrate calpains, remained unsuccessful. We suggest that the Drosophila enzyme is a novel calpain II-like protease.

Original languageEnglish
Pages (from-to)8201-8206
Number of pages6
JournalBiochemistry
Volume31
Issue number35
Publication statusPublished - 1992

Fingerprint

Calpain
Drosophila melanogaster
Sulfhydryl Compounds
Purification
Peptide Hydrolases
Sepharose
Enzymes
Vertebrates
Silver Staining
Divalent Cations
Enzyme Inhibitors
Silver
Application programs
Drosophila
Polyacrylamide Gel Electrophoresis
Ions
Peptides

ASJC Scopus subject areas

  • Biochemistry

Cite this

Purification and characterization of a Ca2+-activated thiol protease from Drosophila melanogaster. / Pintér, Marianna; Friedrich, P.; Friedrich, Peter.

In: Biochemistry, Vol. 31, No. 35, 1992, p. 8201-8206.

Research output: Contribution to journalArticle

Pintér, Marianna ; Friedrich, P. ; Friedrich, Peter. / Purification and characterization of a Ca2+-activated thiol protease from Drosophila melanogaster. In: Biochemistry. 1992 ; Vol. 31, No. 35. pp. 8201-8206.
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