Author:
Dorrazehi Gol Mohammad,Winkle Matthias,Desmet Martin,Stroobant Vincent,Tanriver Gamze,Degand Hervé,Evrard Damien,Desguin Benoît,Morsomme Pierre,Biboy Jacob,Gray Joe,Mitusińska Karolina,Góra Artur,Vollmer Waldemar,Soumillion Patrice
Abstract
AbstractPenicillin binding proteins (PBPs) are involved in biosynthesis, remodeling and recycling of peptidoglycan (PG) in bacteria. PBP-A from Thermosynechococcus elongatus belongs to a cyanobacterial family of enzymes sharing close structural and phylogenetic proximity to class A β-lactamases. With the long-term aim of converting PBP-A into a β-lactamase by directed evolution, we simulated what may happen when an organism like Escherichia coli acquires such a new PBP and observed growth defect associated with the enzyme activity. To further explore the molecular origins of this harmful effect, we decided to characterize deeper the activity of PBP-A both in vitro and in vivo. We found that PBP-A is an enzyme endowed with dd-carboxypeptidase and dd-endopeptidase activities, featuring high specificity towards muropeptides amidated on the d-iso-glutamyl residue. We also show that a low promiscuous activity on non-amidated peptidoglycan deteriorates E. coli’s envelope, which is much higher under acidic conditions where substrate discrimination is mitigated. Besides expanding our knowledge of the biochemical activity of PBP-A, this work also highlights that promiscuity may depend on environmental conditions and how it may hinder rather than promote enzyme evolution in nature or in the laboratory.
Funder
H2020 Marie Skłodowska-Curie Actions
Biotechnology and Biological Sciences Research Council
Publisher
Springer Science and Business Media LLC