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Micellar electrokinetic chromatography: a convenient alternative to colorimetric and high performance liquid chromatographic detection to monitor protease activity.
Electrophoresis. 1998 Sep; 19(12):2083-9.E

Abstract

High performance capillary electrophoresis (HPCE) has been exploited as an analytical method alternative to current procedures for the determination of proteolytic activity of elastases from different sources. Due to some drawbacks with capillary zone electrophoresis (CZE), the mode of operation employed for the assay of elastolytic activity was micellar electrokinetic chromatography (MEKC). Using a background electrolyte consisting of 35 mM sodium tetraborate, pH 9.3, containing 65 mM SDS and 15% v/v methanol, separation of intact peptide substrate from products of proteolytic reaction was easily achieved in a fused-silica capillary of 50 cm effective length x 75 microm ID. This allowed us to determine the rate of hydrolysis of substrates and to calculate the kinetic parameters Km and k(cat) of the proteases investigated. A comparison of these data with those obtained from high performance liquid chromatography (HPLC)-based experiments showed that MEKC is a convenient technique for studying protease kinetics.

Authors+Show Affiliations

Viglio S
Dipartimento di Biochimica A. Castellani, Università di Pavia, Italy.
Zanaboni G
No affiliation info available
Luisetti M
No affiliation info available
Cetta G
No affiliation info available
Guglielminetti M
No affiliation info available
Iadarola P
No affiliation info available

MeSH

Aspergillus fumigatusBoratesChromatography, High Pressure LiquidChromatography, Micellar Electrokinetic CapillaryColorimetryEndopeptidasesHydrogen-Ion ConcentrationKineticsPancreatic ElastasePeptide FragmentsSensitivity and Specificity

Pub Type(s)

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

Language

eng

PubMed ID

9761185

Citation

Viglio, S, et al. "Micellar Electrokinetic Chromatography: a Convenient Alternative to Colorimetric and High Performance Liquid Chromatographic Detection to Monitor Protease Activity." Electrophoresis, vol. 19, no. 12, 1998, pp. 2083-9.
Viglio S, Zanaboni G, Luisetti M, et al. Micellar electrokinetic chromatography: a convenient alternative to colorimetric and high performance liquid chromatographic detection to monitor protease activity. Electrophoresis. 1998;19(12):2083-9.
Viglio, S., Zanaboni, G., Luisetti, M., Cetta, G., Guglielminetti, M., & Iadarola, P. (1998). Micellar electrokinetic chromatography: a convenient alternative to colorimetric and high performance liquid chromatographic detection to monitor protease activity. Electrophoresis, 19(12), 2083-9.
Viglio S, et al. Micellar Electrokinetic Chromatography: a Convenient Alternative to Colorimetric and High Performance Liquid Chromatographic Detection to Monitor Protease Activity. Electrophoresis. 1998;19(12):2083-9. PubMed PMID: 9761185.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Micellar electrokinetic chromatography: a convenient alternative to colorimetric and high performance liquid chromatographic detection to monitor protease activity. AU - Viglio,S, AU - Zanaboni,G, AU - Luisetti,M, AU - Cetta,G, AU - Guglielminetti,M, AU - Iadarola,P, PY - 1998/10/7/pubmed PY - 1998/10/7/medline PY - 1998/10/7/entrez SP - 2083 EP - 9 JF - Electrophoresis JO - Electrophoresis VL - 19 IS - 12 N2 - High performance capillary electrophoresis (HPCE) has been exploited as an analytical method alternative to current procedures for the determination of proteolytic activity of elastases from different sources. Due to some drawbacks with capillary zone electrophoresis (CZE), the mode of operation employed for the assay of elastolytic activity was micellar electrokinetic chromatography (MEKC). Using a background electrolyte consisting of 35 mM sodium tetraborate, pH 9.3, containing 65 mM SDS and 15% v/v methanol, separation of intact peptide substrate from products of proteolytic reaction was easily achieved in a fused-silica capillary of 50 cm effective length x 75 microm ID. This allowed us to determine the rate of hydrolysis of substrates and to calculate the kinetic parameters Km and k(cat) of the proteases investigated. A comparison of these data with those obtained from high performance liquid chromatography (HPLC)-based experiments showed that MEKC is a convenient technique for studying protease kinetics. SN - 0173-0835 UR - https://www.unboundmedicine.com/medline/citation/9761185/Micellar_electrokinetic_chromatography:_a_convenient_alternative_to_colorimetric_and_high_performance_liquid_chromatographic_detection_to_monitor_protease_activity_ L2 - https://doi.org/10.1002/elps.1150191207 DB - PRIME DP - Unbound Medicine ER -