Tags

Type your tag names separated by a space and hit enter

Differences in midgut serine proteinases from larvae of the bruchid beetles Callosobruchus maculatus and Zabrotes subfasciatus.
Arch Insect Biochem Physiol. 2001 May; 47(1):18-28.AI

Abstract

Proteinase activities in the larval midguts of the bruchids Callosobruchus maculatus and Zabrotes subfasciatus were investigated. Both midgut homogenates showed a slightly acidic to neutral pH optima for the hydrolysis of fluorogenic substrates. Proteolysis of epsilon-aminocaproil-Leu-Cys(SBzl)-MCA was totally inhibited by the cysteine proteinase inhibitors E-64 and leupeptin, and was activated by 1.5 mM DTT in both insects, while hydrolysis of the substrate Z-ArgArg-MCA was inhibited by aprotinin and E-64, which suggests that it is being hydrolysed by serine and cysteine proteinases. Gel assays showed that the proteolytic activity in larval midgut of C. maculatus was due to five major cysteine proteinases. However, based on the pattern of E-64 and aprotinin inhibition, proteolytic activity in larval midgut of Z. subfasciatus was not due only to cysteine proteinases. Fractionation of the larval midgut homogenates of both bruchids through ion-exchange chromatography (DEAE-Sepharose) revealed two peaks of activity against Z-ArgArg-MCA for both bruchid species. The fractions from C. maculatus have characteristics of cysteine proteinases, while Z. subfasciatus has one non-retained peak of activity containing cysteine proteinases and another eluted in a gradient of 250-350 mM NaCl. The proteolytic activity of the retained peak is higher at pH 8.8 than at pH 6.0 and corresponds with a single peak that is active against N-p-tosyl-GlyGlyArg-MCA, and sensitive to 250 microM aprotinin (90% inhibition). The peak contains a serine proteinase which hydrolyzes alpha-amylase inhibitor 1 from the common bean (Phaseolus vulgaris). Arch.

Authors+Show Affiliations

Laboratório de Química e Função de Proteínas e Peptídeos, Centro de Biociências e Biotecnologia, Universidade Estadual do Norte Fluminense, Campos dos Goytacazes, Brasil. capsilva@cbb.uenf.brNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

11317332

Citation

Silva, C P., et al. "Differences in Midgut Serine Proteinases From Larvae of the Bruchid Beetles Callosobruchus Maculatus and Zabrotes Subfasciatus." Archives of Insect Biochemistry and Physiology, vol. 47, no. 1, 2001, pp. 18-28.
Silva CP, Terra WR, Lima RM. Differences in midgut serine proteinases from larvae of the bruchid beetles Callosobruchus maculatus and Zabrotes subfasciatus. Arch Insect Biochem Physiol. 2001;47(1):18-28.
Silva, C. P., Terra, W. R., & Lima, R. M. (2001). Differences in midgut serine proteinases from larvae of the bruchid beetles Callosobruchus maculatus and Zabrotes subfasciatus. Archives of Insect Biochemistry and Physiology, 47(1), 18-28.
Silva CP, Terra WR, Lima RM. Differences in Midgut Serine Proteinases From Larvae of the Bruchid Beetles Callosobruchus Maculatus and Zabrotes Subfasciatus. Arch Insect Biochem Physiol. 2001;47(1):18-28. PubMed PMID: 11317332.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Differences in midgut serine proteinases from larvae of the bruchid beetles Callosobruchus maculatus and Zabrotes subfasciatus. AU - Silva,C P, AU - Terra,W R, AU - Lima,R M, PY - 2001/4/24/pubmed PY - 2001/7/20/medline PY - 2001/4/24/entrez SP - 18 EP - 28 JF - Archives of insect biochemistry and physiology JO - Arch Insect Biochem Physiol VL - 47 IS - 1 N2 - Proteinase activities in the larval midguts of the bruchids Callosobruchus maculatus and Zabrotes subfasciatus were investigated. Both midgut homogenates showed a slightly acidic to neutral pH optima for the hydrolysis of fluorogenic substrates. Proteolysis of epsilon-aminocaproil-Leu-Cys(SBzl)-MCA was totally inhibited by the cysteine proteinase inhibitors E-64 and leupeptin, and was activated by 1.5 mM DTT in both insects, while hydrolysis of the substrate Z-ArgArg-MCA was inhibited by aprotinin and E-64, which suggests that it is being hydrolysed by serine and cysteine proteinases. Gel assays showed that the proteolytic activity in larval midgut of C. maculatus was due to five major cysteine proteinases. However, based on the pattern of E-64 and aprotinin inhibition, proteolytic activity in larval midgut of Z. subfasciatus was not due only to cysteine proteinases. Fractionation of the larval midgut homogenates of both bruchids through ion-exchange chromatography (DEAE-Sepharose) revealed two peaks of activity against Z-ArgArg-MCA for both bruchid species. The fractions from C. maculatus have characteristics of cysteine proteinases, while Z. subfasciatus has one non-retained peak of activity containing cysteine proteinases and another eluted in a gradient of 250-350 mM NaCl. The proteolytic activity of the retained peak is higher at pH 8.8 than at pH 6.0 and corresponds with a single peak that is active against N-p-tosyl-GlyGlyArg-MCA, and sensitive to 250 microM aprotinin (90% inhibition). The peak contains a serine proteinase which hydrolyzes alpha-amylase inhibitor 1 from the common bean (Phaseolus vulgaris). Arch. SN - 0739-4462 UR - https://www.unboundmedicine.com/medline/citation/11317332/Differences_in_midgut_serine_proteinases_from_larvae_of_the_bruchid_beetles_Callosobruchus_maculatus_and_Zabrotes_subfasciatus_ L2 - https://doi.org/10.1002/arch.1031 DB - PRIME DP - Unbound Medicine ER -