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Journal of Bacteriology, June 1999, p. 3695-3704, Vol. 181, No. 12
Department of Food Engineering and
Biotechnology, Technion-Israel Institute of Technology, Haifa 32000, Israel,1 and Department of Molecular
Biology and Microbiology, Tufts University, Boston, Massachusetts
021112
Received 4 February 1999/Accepted 13 April 1999
A
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
The Glucuronic Acid Utilization Gene Cluster
from Bacillus stearothermophilus T-6

-EMBL3 genomic library of Bacillus
stearothermophilus T-6 was screened for hemicellulolytic
activities, and five independent clones exhibiting
-xylosidase
activity were isolated. The clones overlap each other and together
represent a 23.5-kb chromosomal segment. The segment contains a cluster
of xylan utilization genes, which are organized in at least three
transcriptional units. These include the gene for the extracellular
xylanase, xylanase T-6; part of an operon coding for an intracellular
xylanase and a
-xylosidase; and a putative 15.5-kb-long
transcriptional unit, consisting of 12 genes involved in the
utilization of
-D-glucuronic acid (GlcUA). The first
four genes in the potential GlcUA operon (orf1,
-2, -3, and -4) code for a putative
sugar transport system with characteristic components of the
binding-protein-dependent transport systems. The most likely natural
substrate for this transport system is aldotetraouronic acid
[2-O-
-(4-O-methyl-
-D-glucuronosyl)-xylotriose] (MeGlcUAXyl3). The following two genes code for an
intracellular
-glucuronidase (aguA) and a
-xylosidase
(xynB). Five more genes (kdgK,
kdgA, uxaC, uxuA, and
uxuB) encode proteins that are homologous to enzymes
involved in galacturonate and glucuronate catabolism. The gene cluster
also includes a potential regulatory gene, uxuR, the
product of which resembles repressors of the GntR family. The apparent
transcriptional start point of the cluster was determined by primer
extension analysis and is located 349 bp from the initial ATG codon.
The potential operator site is a perfect 12-bp inverted repeat located
downstream from the promoter between nucleotides +170 and +181. Gel
retardation assays indicated that UxuR binds specifically to this
sequence and that this binding is efficiently prevented in vitro by
MeGlcUAXyl3, the most likely molecular inducer.
*
Corresponding author. Mailing address: Department of
Food Engineering & Biotechnology, Technion, Haifa 32000, Israel. Phone: 972-4-8293072. Fax: 972-4-8320742. E-mail:
yshoham{at}tx.technion.ac.il.
Present address: Department of Biochemistry, Israel Institute for
Biological Research, Ness-Ziona 70400, Israel.
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