Author: Kranjc, Luka ; Paš, Maja ; Magdevska, Vasilka ; Šnajder, Marko ; Petković, Hrvoje ; Carrillo Rincón, Andrés Felipe ; Poklar Ulrih, Nataša ; Juntes, Polona ; Bahun, Miha

AbstractBackgroundThe thermostable serine protease pernisine originates from the hyperthermophilic Archaeaon Aeropyrum pernix and has valuable industrial applications. Due to its properties, A. pernix cannot be cultivated in standard industrial fermentation facilities. Furthermore, pernisine is a demanding target for heterologous expression in mesophilic heterologous hosts due to the relatively complex processing step involved in its activation.ResultsWe achieved production of active extracellular pernisine in a Streptomyces rimosus host through heterologous expression of the codon-optimised gene by applying step-by-step protein engineering approaches. To ensure secretion of fully active enzyme, the srT signal sequence from the S. rimosus protease was fused to pernisine. To promote correct processing and folding of pernisine, the srT functional cleavage site motif was fused directly to the core pernisine sequence, this way omitting the proregion. Comparative biochemical analysis of the wild-type and recombinant pernisine confirmed that the enzyme produced by S. rimosus retained all of the desired properties of native pernisine. Importantly, the recombinant pernisine also degraded cellular and infectious bovine prion proteins, which is one of the particular applications of this protease.ConclusionFunctional pernisine that retains all of the advantageous properties of the native enzyme from the thermophilic host was successfully produced in a S. rimosus heterologous host. Importantly, we achieved extracellular production of active pernisine, which significantly simplifies further downstream procedures and also omits the need for any pre-processing step for its activation. We demonstrate that S. rimosus can be used as an attractive host for industrial production of recombinant proteins that originate from thermophilic organisms.

02 May 2013·Genome Announcements

Draft Genome Sequence of the Oxytetracycline-Producing Bacterium Streptomyces rimosus ATCC 10970

Article

Author: Liu, Tze-Tze ; Kirby, Ralph ; Hunter, Iain S. ; Tang, Zhenyu ; Pethick, Florence E. ; Guo, Meijin ; Petkovic, Hrvoje ; Hoskisson, Paul A. ; Herron, Paul R. ; MacFadyen, Alison C. ; Chu, Ju ; Sangal, Vartul ; Kosec, Gregor

ABSTRACT We report the draft genome of Streptomyces rimosus (ATCC 10970), a soil isolate that produces oxytetracycline, a commercially important and clinically useful antibiotic.

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