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Title: Initiation, Elongation, and Termination of Bacterial Cellulose Synthesis

Abstract

Cellulose is the major component of the plant cell wall and composed of β-linked glucose units. Use of cellulose is greatly impacted by its physical properties, which are dominated by the number of individual cellulose strand within each fiber and the average length of each strand. Our work described herein provides a complete mechanism for cellulose synthase accounting for its processivity and mechanism of initiation. Using ionic liquids and gel permeation chromatography, we obtain kinetic constants for initiation, elongation, and termination (release of the cellulose strand from the enzyme) for two bacterial cellulose synthases (Gluconacetobacter hansenii and Rhodobacter sphaeroides). Our results show that initiation of synthesis is primer-independent. After initiation, the enzyme undergoes multiple cycles of elongation until the strand is released. The rate of elongation is much faster than that of steady-state turnover. Elongation requires cyclic addition of glucose (from uridine diphosphate-glucose) and then strand translocation by one glucose unit. Translocations greater than one glucose unit result in termination requiring reinitiation. The rate of the strand release, relative to the rate of elongation, determines the processivity of the enzyme. This mechanism and the measured rate constants were supported by kinetic simulation. With the experimentally determined rate constants, we aremore » able to simulate steady-state kinetics and mimic the size distribution of the product. Thus, our results provide for the first time a mechanism for cellulose synthase that accounts for initiation, elongation, and termination.« less

Authors:
; ; ; ORCiD logo [1]; ORCiD logo
  1. Department of Geological Sciences, University of Texas at El Paso, El Paso, Texas 79968, United States
Publication Date:
Research Org.:
Energy Frontier Research Centers (EFRC) (United States). Center for Lignocellulose Structure and Formation (CLSF); Pennsylvania State Univ., University Park, PA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES)
OSTI Identifier:
1423927
Alternate Identifier(s):
OSTI ID: 1470257; OSTI ID: 1508757
Grant/Contract Number:  
SC0001090
Resource Type:
Published Article
Journal Name:
ACS Omega
Additional Journal Information:
Journal Name: ACS Omega Journal Volume: 3 Journal Issue: 3; Journal ID: ISSN 2470-1343
Publisher:
American Chemical Society (ACS)
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; 37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; biofuels (including algae and biomass); bio-inspired; membrane; carbon sequestration; materials and chemistry by design; synthesis (self-assembly); Biological and Medicinal chemistry; Enzyme kinetics; Theory

Citation Formats

McManus, John B., Yang, Hui, Wilson, Liza, Kubicki, James D., and Tien, Ming. Initiation, Elongation, and Termination of Bacterial Cellulose Synthesis. United States: N. p., 2018. Web. https://doi.org/10.1021/acsomega.7b01808.
McManus, John B., Yang, Hui, Wilson, Liza, Kubicki, James D., & Tien, Ming. Initiation, Elongation, and Termination of Bacterial Cellulose Synthesis. United States. https://doi.org/10.1021/acsomega.7b01808
McManus, John B., Yang, Hui, Wilson, Liza, Kubicki, James D., and Tien, Ming. Tue . "Initiation, Elongation, and Termination of Bacterial Cellulose Synthesis". United States. https://doi.org/10.1021/acsomega.7b01808.
@article{osti_1423927,
title = {Initiation, Elongation, and Termination of Bacterial Cellulose Synthesis},
author = {McManus, John B. and Yang, Hui and Wilson, Liza and Kubicki, James D. and Tien, Ming},
abstractNote = {Cellulose is the major component of the plant cell wall and composed of β-linked glucose units. Use of cellulose is greatly impacted by its physical properties, which are dominated by the number of individual cellulose strand within each fiber and the average length of each strand. Our work described herein provides a complete mechanism for cellulose synthase accounting for its processivity and mechanism of initiation. Using ionic liquids and gel permeation chromatography, we obtain kinetic constants for initiation, elongation, and termination (release of the cellulose strand from the enzyme) for two bacterial cellulose synthases (Gluconacetobacter hansenii and Rhodobacter sphaeroides). Our results show that initiation of synthesis is primer-independent. After initiation, the enzyme undergoes multiple cycles of elongation until the strand is released. The rate of elongation is much faster than that of steady-state turnover. Elongation requires cyclic addition of glucose (from uridine diphosphate-glucose) and then strand translocation by one glucose unit. Translocations greater than one glucose unit result in termination requiring reinitiation. The rate of the strand release, relative to the rate of elongation, determines the processivity of the enzyme. This mechanism and the measured rate constants were supported by kinetic simulation. With the experimentally determined rate constants, we are able to simulate steady-state kinetics and mimic the size distribution of the product. Thus, our results provide for the first time a mechanism for cellulose synthase that accounts for initiation, elongation, and termination.},
doi = {10.1021/acsomega.7b01808},
journal = {ACS Omega},
number = 3,
volume = 3,
place = {United States},
year = {2018},
month = {3}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1021/acsomega.7b01808

Citation Metrics:
Cited by: 4 works
Citation information provided by
Web of Science

Figures / Tables:

Figure 1 Figure 1: GPC elution profiles of cellulose tricarbanilates. Cellulose samples (as labeled in the figure) were carbanilated, as described in Materials and Methods. For cellulose synthesized from BcsA-BcsB or AcsA-AcsB, 0.5 μM enzyme with 5 mM UDP-Glc in a reaction volume of 1.0 mL was used, as described in Materialsmore » and Methods. Reactions were allowed to proceed for 12 h. A KD-806M column was used with tetrahydrofuran as the mobile phase at a flow rate of 0.5 mL min−1. The figure shows the molecular weight distribution, converted to DOP on the right axis, above the associated refractive index detection signal.« less

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    Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.