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Title: Site-directed mutagenesis of bacterial cellulose synthase highlights sulfur–arene interaction as key to catalysis
Authors: Sun, Shi-jing
Horikawa, Yoshiki
Wada, Masahisa  kyouindb  KAKEN_id  orcid https://orcid.org/0000-0002-3508-3307 (unconfirmed)
Sugiyama, Junji  kyouindb  KAKEN_id  orcid https://orcid.org/0000-0002-5388-4925 (unconfirmed)
Imai, Tomoya  kyouindb  KAKEN_id  orcid https://orcid.org/0000-0001-7026-0287 (unconfirmed)
Author's alias: 和田, 昌久
杉山, 淳司
今井, 友也
Keywords: Cellulose synthase
CesA
Site-directed mutagenesis
Gluconacetobacter xylinus
Sulfur–arene interaction
Issue Date: 3-Nov-2016
Publisher: Elsevier B.V.
Journal title: Carbohydrate Research
Volume: 434
Start page: 99
End page: 106
Abstract: Cellulose is one of the most abundant biological polymers on Earth, and is synthesized by the cellulose synthase complex in cell membranes. Although many cellulose synthase genes have been identified over the past 25 years, functional studies of cellulose synthase using recombinant proteins have rarely been conducted. In this study, we conducted a functional analysis of cellulose synthase with site-directed mutagenesis, by using recombinant cellulose synthase reconstituted in living Escherichia coli cells that we recently constructed (cellulose-synthesizing E. coli, CESEC). We demonstrated that inactivating mutations at an important amino acid residue reduced cellulose production. In this study, an interesting loss-of-function mutation occurred on Cys308, whose main chain carbonyl plays an important role for locating the cellulose terminus. Mutating this cysteine to serine, thus changing sulfur to oxygen in the side chain, abolished cellulose production in addition to other apparent detrimental mutations. This unexpected result highlights that the thiol side-chain of this cysteine plays an active role in catalysis, and additional mutation experiments indicated that the sulfur–arene interaction around Cys308 is a key in cellulose-synthesizing activity. Data obtained by CESEC shed light on the function of cellulose synthase in living cells, and will deepen our understanding of the mechanism of cellulose synthase.
Rights: © 2016 The Authors. Published by Elsevier Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
URI: http://hdl.handle.net/2433/217352
DOI(Published Version): 10.1016/j.carres.2016.08.009
PubMed ID: 27623440
Appears in Collections:Journal Articles

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