Abstract
The increased exploitation of microbial sequencing methods has shed light on the high diversity of new microorganisms named Candidate Phyla Radiation (CPR). CPR are mainly detected via 16S rRNA/metabarcoding analyses or metagenomics and are found to be abundant in all environments and present in different human microbiomes. These microbes, characterized by their symbiotic/epiparasitic lifestyle with bacteria, are directly exposed to competition with other microorganisms sharing the same ecological niche. Recently, a rich repertoire of enzymes with antibiotic resistance activity has been found in CPR genomes by using an in silico adapted screening strategy. This reservoir has shown a high prevalence of putative beta-lactamase-encoding genes. We expressed and purified five putative beta-lactamase sequences having the essential domains and functional motifs from class A and class B beta-lactamase. Their enzymatic activities were tested against various beta-lactam substrates using liquid chromatography-mass spectrometry (LC-MS) and showed some beta-lactamase activity even in the presence of a beta-lactamase inhibitor. In addition, ribonuclease activity was demonstrated against RNA that was not inhibited by sulbactam and EDTA. None of these proteins could degrade single- and double-stranded-DNA. This study is the first to express and test putative CPR beta-lactamase protein sequences in vitro. Our findings highlight that the reduced genomes of CPR members harbor sequences encoding for beta-lactamases known to be multifunction hydrolase enzymes.