Expression of Streptomyces genes encoding extracellular enzymes in Brevibacterium lactofermentum: secretion proceeds by removal of the same leader peptide as in Streptomyces lividans

Cadenas, Rosa F.; Gil, José A.; Martı́n, Juan F.

doi:10.1007/bf00170087

Cited by 15 publications

(5 citation statements)

References 28 publications

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“…The natural substrate spectrum of C. glutamicum further includes sugars like ribose or maltose, alcohols like ethanol or myo ‐inositol and organic acids like acetate, citrate, lactate, propionate and pyruvate and amino acids like l ‐glutamate (Kramer et al ., ; Dominguez et al ., ; Kiefer et al ., ; Gerstmeir et al ., ; Eikmanns, ; Moon et al ., ; Polen et al ., ; Stansen et al ., ; Krings et al ., ; Frunzke et al ., ; Kato et al ., ; Neuner and Heinzle, ). Within the flexible feedstock concept, the substrate spectrum of C. glutamicum has been extended by metabolic engineering to allow access to starch, cellobiose, lactose, galactose and glycerol as well as succinate, fumarate and malate as carbon sources (Brabetz et al ., ; Cadenas et al ., ; Kotrba et al ., ; Barrett et al ., ; Seibold et al ., ; Tateno et al ., ; Rittmann et al ., ; Youn et al ., ; ).…”

Section: Introductionmentioning

confidence: 97%

Accelerated pentose utilization by Corynebacterium glutamicum for accelerated production of lysine, glutamate, ornithine and putrescine

Meiswinkel

Gopinath

Lindner

et al. 2012

Microbial Biotechnology

145

120

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SummaryBecause of their abundance in hemicellulosic wastes arabinose and xylose are an interesting source of carbon for biotechnological production processes. Previous studies have engineered several Corynebacterium glutamicum strains for the utilization of arabinose and xylose, however, with inefficient xylose utilization capabilities. To improve xylose utilization, different xylose isomerase genes were tested in C. glutamicum. The gene originating from Xanthomonas campestris was shown to have the highest effect, resulting in growth rates of 0.14 h−1, followed by genes from Bacillus subtilis, Mycobacterium smegmatis and Escherichia coli. To further increase xylose utilization different xylulokinase genes were expressed combined with X. campestris xylose isomerase gene. All combinations further increased growth rates of the recombinant strains up to 0.20 h−1 and moreover increased biomass yields. The gene combination of X. campestris xylose isomerase and C. glutamicum xylulokinase was the fastest growing on xylose and compared with the previously described strain solely expressing E. coli xylose isomerase gene delivered a doubled growth rate. Productivity of the amino acids glutamate, lysine and ornithine, as well as the diamine putrescine was increased as well as final titres except for lysine where titres remained unchanged. Also productivity in medium containing rice straw hydrolysate as carbon source was increased.Funding Information No funding information provided.

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Section: Introductionmentioning

confidence: 97%

Accelerated pentose utilization by Corynebacterium glutamicum for accelerated production of lysine, glutamate, ornithine and putrescine

Meiswinkel

Gopinath

Lindner

et al. 2012

Microbial Biotechnology

145

120

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“…In our laboratory, several antibiotic resistance genes have been successfully used for developing a pBL1-based cloning system for B. lactofermentum: the kanamycin resistance gene (kan) from TnS, the hygromycin resistance gene (hyg) from Streptomyces hygroscopicus, and the chloramphenicol resistance gene (cat) from Streptomyces acrimycini (20,21). Cadenas et al (2) described the efficient expression in B. lactofermentum in pBL1-derived vectors of a heterologous gene coding for an a.-amylase from Streptomyces griseus. Other groups have described the use of Escherichia coli and Bacillus subtilis phenotypic markers in corynebacteria (14,16,22).…”

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confidence: 99%

Characterization of a region of plasmid pBL1 of Brevibacterium lactofermentum involved in replication via the rolling circle model

et al. 1994

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The minimal region for autonomous replication of pBL1, a 4.5-kb cryptic plasmid of Brevibacterium lactofermentum ATCC 13869 that has been used to construct a variety of corynebacterium vectors, was shown to be contained on a 1.8-kb HindII-SphI DNA fragment. This region contains two open reading frames (ORFs) (ORF1 and ORF5) which are essential for pBL1 replication in B. lactofermentum. Accumulation of single-strand intermediates in some of the constructions indicates that plasmid pBL1 replicates via the rolling circle replication model; its plus strand and minus strand were identified by hybridization with two synthetic oligonucleotide probes complementary to each pBL1 strand. ORF1 seems to encode the Rep protein and showed partial homology with sequences for Rep proteins from Streptomyces plasmids which replicate via rolling circle replication such as pUJ101, pSB24, and pJVl.pBL1 is a multicopy plasmid isolated from Brevibacterium lactofermentum ATCC 13869 (19) and later described as pAM330 (14) Streptomyces lividans. pUL61 was unstable in B. lactofermentum, and two stable deletion derivatives (pUL330 and pUL340) appeared after B. lactofermentum had been transformed with pUL61. A similar instability has also been observed in most other small plasmids from gram-positive bacteria. The main characteristic of plasmids showing instability is their mode of replication via the rolling circle replication (RCR) mechanism. RCR plasmids were classified into four families according to homologies of their replication proteins (Rep) and the position of the double-stranded origin (DSO) (7). RCR plasmids have the information to synthesize their own Rep protein. The Rep protein initiates replication by nicking the DNA at the DSO sequence, after which replication of the plus strand begins. The newly generated single-stranded DNA (ssDNA) intermediates (plus strand) serve as a template for minus strand synthesis. The conversion of the ssDNA intermediates to duplex forms requires the recognition of another sequence, different and separated from the DSO sequence, called single-stranded origin (SSO). The absence or nonfunctionality of SSO results in the accumulation of ssDNA. The SSO sequence is rather specific, and it is usually recognized inefficiently in hosts different from the parental one (7, 15).We describe in this paper the identification and characterization of the region involved in replication functions of pBL1. Our results indicate that pBL1 belongs to the group of plasmids which replicate via the RCR mechanism. MATERIALS AND METHODSPlasmids, bacterial strains, and transformation conditions. Plasmids used in this study are listed in Table 1. B. lactofermentum BL31 (20), a pBL1-cured strain, and C. glutamicum ATCC 13032 were grown in Trypticase soy broth (TSB) medium and transformed by electroporation as described by Dunican and Shivnan (5). E. coli DH5ao was grown in L broth and transformed by the method of Hanahan (8).

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“…In species of Streptomyces the processing of ␣-amylase is known to be mediated by intracellular specific proteolytic cleavage (10). The same proteolytic cleavage occurs in N. lactamdurans and also in corynebacteria (5). The amount of extracellular ␣-amylase is an indication of the level of expression of the reporter gene and can be used to compare different promoters.…”

Section: Discussionmentioning

confidence: 99%

“…During studies on protein secretion in S. lividans (10,23) and corynebacteria (5) using the amy gene of Streptomyces griseus, we found that the amount of ␣-amylase secreted was proportional to the strength of the promoter used to express the amy gene since there is no bottleneck in amylase secretion up to very high levels of expression of the amy gene (24). Processing of the enzyme during secretion occurred by releasing the same leader peptide in both corynebacteria and Streptomyces species (5). Later, we observed that the S. griseus amy gene can be used as a marker for detection of N. lactamdurans transformants (15).…”

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confidence: 99%

Amy as a reporter gene for promoter activity in Nocardia lactamdurans: comparison of promoters of the cephamycin cluster

Chary

Fuente

Liras

et al. 1997

Appl Environ Microbiol

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Promoter probe vectors containing the pA origin of replication and the Streptomyces griseus promoterless amy gene (encoding ␣-amylase) as reporter have been constructed to study transcription initiation regions in Nocardia lactamdurans. In some of the promoter probe vectors the phage fd terminator has been introduced to avoid readthrough expression from upstream sequences. By using these vectors, four different transcription initiation regions of the cephamycin gene cluster have been studied in N. lactamdurans. The bla gene encoding a ␤-lactamase has a relatively strong promoter. Two other separate promoters corresponding to the lat and cefD genes (encoding, respectively, lysine-6-aminotransferase and isopenicillin N-epimerase) showed weak transcription initiation ability. These two promoters are arranged in a bidirectional transcription initiation region located in the center of the cephamycin gene cluster. The cmcH gene (encoding 3-hydroxymethylcephem carbamoyltransferase) upstream region did not contain a functional promoter, suggesting that cmcH is transcribed as a part of a polycistronic mRNA. The native amy promoter is used very efficiently in N. lactamdurans, resulting in secretion of high levels of extracellular ␣-amylase.

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Expression of Streptomyces genes encoding extracellular enzymes in Brevibacterium lactofermentum: secretion proceeds by removal of the same leader peptide as in Streptomyces lividans

Cited by 15 publications

References 28 publications

Accelerated pentose utilization by Corynebacterium glutamicum for accelerated production of lysine, glutamate, ornithine and putrescine

Accelerated pentose utilization by Corynebacterium glutamicum for accelerated production of lysine, glutamate, ornithine and putrescine

Characterization of a region of plasmid pBL1 of Brevibacterium lactofermentum involved in replication via the rolling circle model

Amy as a reporter gene for promoter activity in Nocardia lactamdurans: comparison of promoters of the cephamycin cluster

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