Dataset Information

Structure and function of a novel GH8 endoglucanase from the bacterial cellulose synthase complex of Raoultella ornithinolytica.

ABSTRACT: Cellulose synthesis in bacteria is a complex process involving the concerted action of several enzymes whose genes are often organized in operons. This process influences many fundamental physiological aspects such as bacteria and host interaction, biofilm formation, among others. Although it might sound contradictory, the participation of cellulose-degrading enzymes is critical to this process. The presence of endoglucanases from family 8 of glycosyl hydrolases (GH8) in bacterial cellulose synthase (Bcs) complex has been described in different bacteria, including the model organism Komagataeibacter xylinus; however, their role in this process is not completely understood. In this study, we describe the biochemical characterization and three-dimensional structure of a novel GH8 member from Raoultella ornithinolytica, named AfmE1, which was previously identified by our group from the metagenomic analysis of the giant snail Achatina fulica. Our results demonstrated that AfmE1 is an endo-?-1,4-glucanase, with maximum activity in acidic to neutral pH over a wide temperature range. This enzyme cleaves cello-oligosaccharides with a degree of polymerization ? 5 and presents six glucosyl-binding subsites. The structural comparison of AfmE1 with other GH8 endoglucanases showed significant structural dissimilarities in the catalytic cleft, particularly in the subsite +3, which correlate with different functional mechanisms, such as the recognition of substrate molecules having different arrangements and crystallinities. Together, these findings provide new insights into molecular and structural features of evolutionarily conserved endoglucanases from the bacterial cellulose biosynthetic machinery.

SUBMITTER: Scapin SMN

PROVIDER: S-EPMC5407803 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structure and function of a novel GH8 endoglucanase from the bacterial cellulose synthase complex of Raoultella ornithinolytica.

Scapin Sandra Mara Naressi SMN Souza Flavio Henrique Moreira FHM Zanphorlin Leticia Maria LM de Almeida Thamyres Silva TS Sade Youssef Bacila YB Cardoso Alexander Machado AM Pinheiro Guilherme Luiz GL Murakami Mario Tyago MT

PloS one 20170427 4

Cellulose synthesis in bacteria is a complex process involving the concerted action of several enzymes whose genes are often organized in operons. This process influences many fundamental physiological aspects such as bacteria and host interaction, biofilm formation, among others. Although it might sound contradictory, the participation of cellulose-degrading enzymes is critical to this process. The presence of endoglucanases from family 8 of glycosyl hydrolases (GH8) in bacterial cellulose synt ...[more]

PMID: 28448629

Similar Datasets

Project description:The processive cycle of the bacterial cellulose synthase (Bcs) includes the addition of a single glucose moiety to the end of a growing cellulose chain followed by the translocation of the nascent chain across the plasma membrane. The mechanism of this translocation and its precise location within the processive cycle are not well understood. In particular, the molecular details of how a polymer (cellulose) whose basic structural unit is a dimer (cellobiose) can be constructed by adding one monomer (glucose) at a time are yet to be elucidated. Here, we have utilized molecular dynamics simulations and free energy calculations to the shed light on these questions. We find that translocation forward by one glucose unit is quite favorable energetically, giving a free energy stabilization of greater than 10 kcal/mol. In addition, there is only a small barrier to translocation, implying that translocation is not rate limiting within the Bcs processive cycle (given experimental rates for cellulose synthesis in vitro). Perhaps most significantly, our results also indicate that steric constraints at the transmembrane tunnel entrance regulate the dimeric structure of cellulose. Namely, when a glucose molecule is added to the cellulose chain in the same orientation as the acceptor glucose, the terminal glucose freely rotates upon forward motion, thus suggesting a regulatory mechanism for the dimeric structure of cellulose. We characterize both the conserved and non-conserved enzyme-polysaccharide interactions that drive translocation, and find that 20 of the 25 residues that strongly interact with the translocating cellulose chain in the simulations are well conserved, mostly with polar or aromatic side chains. Our results also allow for a dynamical analysis of the role of the so-called `finger helix' in cellulose translocation that has been observed structurally. Taken together, these findings aid in the elucidation of the translocation steps of the Bcs processive cycle and may be widely relevant to polysaccharide synthesizing or degrading enzymes that couple catalysis with chain translocation.

Dataset Information

Structure and function of a novel GH8 endoglucanase from the bacterial cellulose synthase complex of Raoultella ornithinolytica.

Publications

Structure and function of a novel GH8 endoglucanase from the bacterial cellulose synthase complex of Raoultella ornithinolytica.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets