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Applied and Environmental Microbiology, September 2001, p. 4166-4176, Vol. 67, No. 9
Instituto de Biología Molecular y Celular de
Rosario (IBR-CONICET) and Departamento de Microbiología,
Facultad de Ciencias Bioquímicas y Farmacéuticas,
Universidad Nacional de Rosario, Suipacha 531, 2000-Rosario,
Argentina,1 and Department of
Molecular Microbiology, John Innes Centre, Colney Lane, Norwich NR4
7UH, United Kingdom2
Received 5 March 2001/Accepted 27 June 2001
Two genes, accB and accE, that form
part of the same operon, were cloned from Streptomyces
coelicolor A3(2). AccB is homologous to the carboxyl
transferase domain of several propionyl coezyme A (CoA) carboxylases
and acyl-CoA carboxylases (ACCases) of actinomycete origin, while AccE
shows no significant homology to any known protein. Expression of
accB and accE in Escherichia
coli and subsequent in vitro reconstitution of enzyme activity
in the presence of the biotinylated protein AccA1 or AccA2 confirmed
that AccB was the carboxyl transferase subunit of an ACCase.
The additional presence of AccE considerably enhanced the activity of
the enzyme complex, suggesting that this small polypeptide is a
functional component of the ACCase. The impossibility of
obtaining an accB null mutant and the thiostrepton
growth dependency of a tipAp accB conditional mutant
confirmed that AccB is essential for S. coelicolor
viability. Normal growth phenotype in the absence of the inducer was
restored in the conditional mutant by the addition of exogenous
long-chain fatty acids in the medium, indicating that the
inducer-dependent phenotype was specifically related to a conditional
block in fatty acid biosynthesis. Thus, AccB, together with AccA2,
which is also an essential protein (E. Rodriguez and H. Gramajo,
Microbiology 143:3109-3119, 1999), are the most likely components of
an ACCase whose main physiological role is the synthesis of
malonyl-CoA, the first committed step of fatty acid synthesis. Although
normal growth of the conditional mutant was restored by fatty acids,
the cultures did not produce actinorhodin or undecylprodigiosin,
suggesting a direct participation of this enzyme complex in the supply
of malonyl-CoA for the synthesis of these secondary metabolites.
0099-2240/01/$04.00+0 DOI: 10.1128/AEM.67.9.4166-4176.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Role of an Essential Acyl Coenzyme A Carboxylase in the Primary
and Secondary Metabolism of Streptomyces
coelicolor A3(2)
*
Corresponding author. Mailing address: IBR,
Departamento de Microbiología, Facultad de Ciencias
Bioquímicas y Farmacéuticas, Universidad Nacional de
Rosario, Suipacha 531, 2000-Rosario, Argentina. Phone: 54-341-4350661. Fax: 54-341-4390465. E-mail: gramajo{at}infovia.com.ar.
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