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J Bacteriol, February 1998, p. 809-814, Vol. 180, No. 4
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Mutational Biosynthesis of Novel Rapamycins by a Strain of Streptomyces hygroscopicus NRRL 5491 Disrupted in rapL, Encoding a Putative Lysine Cyclodeaminase

Lake Ee Khaw,¹ Günter A. Böhm,² Su Metcalfe,³ James Staunton,² and Peter F. Leadlay^1,*

Cambridge Centre for Molecular Recognition and Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA,¹ Cambridge Centre for Molecular Recognition and University Chemical Laboratory, University of Cambridge, Cambridge CB2 1EW,² and University of Cambridge Clinical School, Department of Surgery, Addenbrooke's Hospital, Cambridge CB2 2QQ,³ United Kingdom

Received 16 September 1997/Accepted 10 December 1997

The gene rapL lies within the region of the Streptomyces hygroscopicus chromosome which contains the biosynthetic gene cluster for the immunosuppressant rapamycin. Introduction of a frameshift mutation into rapL by C31 phage-mediated gene replacement gave rise to a mutant which did not produce significant amounts of rapamycin. Growth of this rapL mutant on media containing added L-pipecolate restored wild-type levels of rapamycin production, consistent with a proposal that rapL encodes a specific L-lysine cyclodeaminase important for the production of the L-pipecolate precursor. In the presence of added proline derivatives, rapL mutants synthesized novel rapamycin analogs, indicating a relaxed substrate specificity for the enzyme catalyzing pipecolate incorporation into the macrocycle.

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^* Corresponding author. Mailing address: Department of Biochemistry, University of Cambridge, 80 Tennis Court Rd., Cambridge CB2 1GA, United Kingdom. Phone: 44-1223-333656. Fax: 44-1223-333656. E-mail: pf110{at}mole.bio.cam.ac.uk.

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