Determination of Km and kcat for signal peptidase I using a full length secretory precursor, pro-OmpA-nuclease A

Autor:	S, Chatterjee, D, Suciu, R E, Dalbey, P C, Kahn, M, Inouye
Rok vydání:	1995
Předmět:	Kinetics Escherichia coli Proteins Hydrolysis Endopeptidases Molecular Sequence Data Serine Endopeptidases Escherichia coli Membrane Proteins Micrococcal Nuclease Amino Acid Sequence Protein Precursors Protein Processing Post-Translational Bacterial Outer Membrane Proteins
Zdroj:	Journal of molecular biology. 245(4)
ISSN:	0022-2836
Popis:	An effective method for the determination of the activity of signal peptidase I (SPase I) of Escherichia coli is established using the hybrid protein pro-OmpA-nuclease A as substrate. Pro-OmpA-nuclease A, a hybrid secretory precursor was purified to homogeneity under denaturing conditions. When this protein was refolded, it could be quantitatively processed by purified SPase I. The Km of signal peptidase I was 0.0165 mM. The kcat was 8.73 s-1. The Km is 50 to 100 times lower than that obtained with peptide substrates indicating that SPase I has a significantly greater affinity for the protein substrate. The turnover number, kcat, is two to four orders of magnitude greater as well. Thus, the specificity constant, kcat/Km is six orders of magnitude greater with pro-OmpA-nuclease A than with peptide substrates. This is the first determination of kinetics of SPase I with a protein substrate.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=pmid________::674842adc340d703cf7102c619771ab2 https://pubmed.ncbi.nlm.nih.gov/7837264 Zobrazit plný text záznamu Full Text from ScienceDirect