6sc4

Publication Abstract from PubMed

Environments previously thought to be uninhabitable offer a tremendous wealth of unexplored microorganisms and enzymes. In this paper, we present the discovery and characterization of a novel gamma-carbonic anhydrase (gamma-CA) from the polyextreme Red Sea brine pool Discovery Deep (2141 m depth, 44.8 degrees C, 26.2% salt) by single-cell genome sequencing. The extensive analysis of the selected gene helps demonstrate the potential of this culture-independent method. The enzyme was expressed in the bioengineered haloarchaeon Halobacterium sp. NRC-1 and characterized by X-ray crystallography and mutagenesis. The 2.6 A crystal structure of the protein shows a trimeric arrangement. Within the gamma-CA, several possible structural determinants responsible for the enzyme's salt stability could be highlighted. Moreover, the amino acid composition on the protein surface and the intra- and intermolecular interactions within the protein differ significantly from those of its close homologs. To gain further insights into the catalytic residues of the gamma-CA enzyme, we created a library of variants around the active site residues and successfully improved the enzyme activity by 17-fold. As several gamma-CAs have been reported without measurable activity, this provides further clues as to critical residues. Our study reveals insights into the halophilic gamma-CA activity and its unique adaptations. The study of the polyextremophilic carbonic anhydrase provides a basis for outlining insights into strategies for salt adaptation, yielding enzymes with industrially valuable properties, and the underlying mechanisms of protein evolution.

Crystal Structure and Active Site Engineering of a Halophilic gamma-Carbonic Anhydrase.,Vogler M, Karan R, Renn D, Vancea A, Vielberg MT, Grotzinger SW, DasSarma P, DasSarma S, Eppinger J, Groll M, Rueping M Front Microbiol. 2020 Apr 28;11:742. doi: 10.3389/fmicb.2020.00742. eCollection, 2020. PMID:32411108^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.