Screening Collagenase Activity in Bacterial Lysate for Directed Enzyme Applications.
bacterial lysate screening
collagenase
directed enzyme evolution
enzymatic assay
molecular dynamics
protein expression
Journal
International journal of molecular sciences
ISSN: 1422-0067
Titre abrégé: Int J Mol Sci
Pays: Switzerland
ID NLM: 101092791
Informations de publication
Date de publication:
09 Aug 2021
09 Aug 2021
Historique:
received:
29
06
2021
revised:
04
08
2021
accepted:
04
08
2021
entrez:
27
8
2021
pubmed:
28
8
2021
medline:
16
9
2021
Statut:
epublish
Résumé
Collagenases are essential enzymes capable of digesting triple-helical collagen under physiological conditions. These enzymes play a key role in diverse physiological and pathophysiological processes. Collagenases are used for diverse biotechnological applications, and it is thus of major interest to identify new enzyme variants with improved characteristics such as expression yield, stability, or activity. The engineering of new enzyme variants often relies on either rational protein design or directed enzyme evolution. The latter includes screening of a large randomized or semirational genetic library, both of which require an assay that enables the identification of improved variants. Moreover, the assay should be tailored for microplates to allow the screening of hundreds or thousands of clones. Herein, we repurposed the previously reported fluorogenic assay using 3,4-dihydroxyphenylacetic acid for the quantitation of collagen, and applied it in the detection of bacterial collagenase activity in bacterial lysates. This enabled the screening of hundreds of
Identifiants
pubmed: 34445258
pii: ijms22168552
doi: 10.3390/ijms22168552
pmc: PMC8395246
pii:
doi:
Substances chimiques
Bacterial Proteins
0
Recombinant Proteins
0
Collagen
9007-34-5
Microbial Collagenase
EC 3.4.24.3
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Références
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