Crystal Structure and Active Site Engineering of a Halophilic γ-Carbonic Anhydrase.
Type
ArticleAuthors
Vogler, Malvina M.
Karan, Ram
Renn, Dominik
Vancea, Alexandra

Vielberg, Marie-Theres
Grötzinger, Stefan W.
DasSarma, Priya
DasSarma, Shiladitya
Eppinger, Jörg

Groll, Michael
Rueping, Magnus

KAUST Department
Academic AffairsBiological & Organometallic Catalysis Laboratories
Biological and Environmental Sciences and Engineering (BESE) Division
Bioscience Program
Chemical Science Program
KAUST Catalysis Center (KCC)
Office of the VP
Physical Science and Engineering (PSE) Division
Date
2020-04-28Submitted Date
2020-02-18Permanent link to this record
http://hdl.handle.net/10754/662868
Metadata
Show full item recordAbstract
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 γ-carbonic anhydrase (γ-CA) from the polyextreme Red Sea brine pool Discovery Deep (2141 m depth, 44.8°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 Å crystal structure of the protein shows a trimeric arrangement. Within the γ-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 γ-CA enzyme, we created a library of variants around the active site residues and successfully improved the enzyme activity by 17-fold. As several γ-CAs have been reported without measurable activity, this provides further clues as to critical residues. Our study reveals insights into the halophilic γ-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.Citation
Vogler, M., Karan, R., Renn, D., Vancea, A., Vielberg, M.-T., Grötzinger, S. W., … Rueping, M. (2020). Crystal Structure and Active Site Engineering of a Halophilic γ-Carbonic Anhydrase. Frontiers in Microbiology, 11. doi:10.3389/fmicb.2020.00742Sponsors
The research reported in this publication was supported by the King Abdullah University of Science and Technology (KAUST), Kingdom of Saudi Arabia. The staff of the Beamline X06SA at the Paul Scherrer Institute, SLS, Villigen, Switzerland, is acknowledged for assistance during data collection. Work in the DasSarma laboratory was supported by grant 80NSSC19K0463.Publisher
Frontiers Media SAJournal
Frontiers in microbiologyPubMed ID
32411108PubMed Central ID
PMC7199487Additional Links
https://www.frontiersin.org/article/10.3389/fmicb.2020.00742/fullhttps://www.frontiersin.org/articles/10.3389/fmicb.2020.00742/pdf
ae974a485f413a2113503eed53cd6c53
10.3389/fmicb.2020.00742
Scopus Count
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