Title: Isolation and characterization of two cellulose morphology mutants of Gluconacetobacter hansenii ATCC23769 producing cellulose with lower crystallinity

Abstract

Gluconacetobacter hansenii, a Gram-negative bacterium, produces and secrets highly crystalline cellulose into growth medium, and has long been used as a model system for studying cellulose synthesis in higher plants. Cellulose synthesis involves the formation of β-1,4 glucan chains via the polymerization of glucose units by a multi-enzyme cellulose synthase complex (CSC). These glucan chains assemble into ordered structures including crystalline microfibrils. AcsA is the catalytic subunit of the cellulose synthase enzymes in the CSC, and AcsC is required for the secretion of cellulose. However, little is known about other proteins required for the assembly of crystalline cellulose. To address this question, we visually examined cellulose pellicles formed in growth media of 763 individual colonies of G. hansenii generated via Tn5 transposon insertion mutagenesis, and identified 85 that produced cellulose with altered morphologies. X-ray diffraction analysis of these 85 mutants identified two that produced cellulose with significantly lower crystallinity than wild type. The gene disrupted in one of these two mutants encoded a lysine decarboxylase and that in the other encoded an alanine racemase. Solid-state NMR analysis revealed that cellulose produced by these two mutants contained increased amounts of non-crystalline cellulose and monosaccharides associated with non-cellulosic polysaccharides as compared tomore » the wild type. Monosaccharide analysis detected higher percentages of galactose and mannose in cellulose produced by both mutants. Field emission scanning electron microscopy showed that cellulose produced by the mutants was unevenly distributed, with some regions appearing to contain deposition of non-cellulosic polysaccharides; however, the width of the ribbon was comparable to that of normal cellulose. As both lysine decarboxylase and alanine racemase are required for the integrity of peptidoglycan, we propose a model for the role of peptidoglycan in the assembly of crystalline cellulose.« less

Authors:

Deng, Ying ^[1]; Nagachar, Nivedita ^[1]; Fang, Lin ^[2]; Luan, Xin ^[1]; Catchmark, Jeffrey M. ^[2]; Tien, Ming ^[1]; Kao, Teh -hui ^[1]; Lai, Hsin -Chih ^[3]

---

+ Show Author Affiliations

1. The Pennsylvania State Univ., University Park, PA (United States)
2. Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States)
3. Chang-Gung Univ. (Taiwan)

Publication Date:

Thu Mar 19 00:00:00 EDT 2015

Research Org.:

Pennsylvania State Univ., University Park, PA (United States); Energy Frontier Research Centers (EFRC) (United States). Center for Lignocellulose Structure and Formation (CLSF)

Sponsoring Org.:

USDOE Office of Science (SC), Basic Energy Sciences (BES)

OSTI Identifier:

1194073

Grant/Contract Number:  

SC0001090

Resource Type:

Accepted Manuscript

Journal Name:

PLoS ONE

Additional Journal Information:

Journal Volume: 10; Journal Issue: 3; Journal ID: ISSN 1932-6203

Publisher:

Public Library of Science

Country of Publication:

United States

Language:

English

Subject:

59 BASIC BIOLOGICAL SCIENCES