Novel thermostable antibiotic resistance enzymes from the Atlantis II Deep Red Sea brine pool

• Published in: Microbial biotechnology Vol. 10; no. 1; pp. 189 - 202
• Main Authors: Elbehery, Ali H. A., Leak, David J., Siam, Rania
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.01.2017; John Wiley and Sons Inc
• Subjects: Aminoglycoside antibiotics; Aminoglycoside phosphotransferase; Antibiotic resistance; Antibiotics; Antimicrobial agents; beta-Lactamases - chemistry; beta-Lactamases - genetics; beta-Lactamases - metabolism; Biochemical characteristics; Biochemistry; Biomedical materials; Blast resistance; Cloning, Molecular; Computational Biology; Disease control; Drug resistance; Drug Resistance, Bacterial; E coli; Enzyme Stability; Enzymes; Evolution; Fluorides; Gene Expression; Geologic Sediments; Identification; Incubation; Indian Ocean; Kanamycin Kinase - chemistry; Kanamycin Kinase - genetics; Kanamycin Kinase - metabolism; Metagenome; Metagenomics; Microorganisms; Open Reading Frames; Phosphotransferase; Polypeptides; Potassium; Saline water; Salinity; Salts; Sequence Analysis, DNA; Sequence Homology; Studies; Temperature; Thermal stability; Red Sea
• ISSN: 1751-7915; 1751-7915
• DOI: 10.1111/1751-7915.12468

Summary The advent of metagenomics has greatly facilitated the discovery of enzymes with useful biochemical characteristics for industrial and biomedical applications, from environmental niches. In this study, we used sequence‐based metagenomics to identify two antibiotic resistance enzymes from the secluded, lower convective layer of Atlantis II Deep Red Sea brine pool (68°C, ~2200 m depth and 250‰ salinity). We assembled > 4 000 000 metagenomic reads, producing 43 555 contigs. Open reading frames (ORFs) called from these contigs were aligned to polypeptides from the Comprehensive Antibiotic Resistance Database using BLASTX. Two ORFs were selected for further analysis. The ORFs putatively coded for 3′‐aminoglycoside phosphotransferase [APH(3′)] and a class A beta‐lactamase (ABL). Both genes were cloned, expressed and characterized for activity and thermal stability. Both enzymes were active in vitro, while only APH(3′) was active in vivo. Interestingly, APH(3′) proved to be thermostable (Tm = 61.7°C and ~40% residual activity after 30 min of incubation at 65°C). On the other hand, ABL was not as thermostable, with a Tm = 43.3°C. In conclusion, we have discovered two novel AR enzymes with potential application as thermophilic selection markers. We used sequence‐based metagenomics to identify extremophilic antibiotic resistance (AR) enzymes from the Atlantis II Deep Red Sea brine pool. Open reading frames putatively coded for 3′‐aminoglycoside phosphotransferase and a class A beta‐lactamase were cloned, expressed and characterized for activity and thermal stability. This study identifies novel AR enzymes, with potential application as thermophilic selection markers.