Abstract:
:The production of candidate affinity proteins in a soluble form, for downstream characterization, is often a time-consuming step in combinatorial protein engineering methods. Here, a novel approach for efficient production of candidate clones is described based on direct cleavage of the affinity protein from the surface of Staphylococcus carnosus, followed by affinity purification. To find a suitable strategy, three new fusion protein constructs were created, introducing a protease site for specific cleavage and purification tags for affinity chromatography purifications into the staphylococcal display vector. The three modified strains were evaluated in terms of transformation frequency, surface expression level and protease cleavage efficiency. A protocol for efficient affinity purification of protease-released affinity proteins using the introduced fusion-tags was successfully used, and the functionality of protease-treated and purified proteins was verified in a biosensor assay. To evaluate the devised method, a previously selected HER2-specific affibody was produced applying the new principle and was used to analyze HER2 expression on human breast cancer cells.
journal_name
FEMS Microbiol Lettjournal_title
FEMS microbiology lettersauthors
Kronqvist N,Löfblom J,Severa D,Ståhl S,Wernérus Hdoi
10.1111/j.1574-6968.2007.00990.xsubject
Has Abstractpub_date
2008-01-01 00:00:00pages
128-36issue
1eissn
0378-1097issn
1574-6968pii
FML990journal_volume
278pub_type
杂志文章abstract::Bifidobacteria are normal inhabitants of the human gut, and members of which are generally considered to be probiotic. Before exerting their beneficial properties, they must survive and persist in the physiological concentrations (0.05-2%) of bile in the gut. In this work, the functional role of tlyC1 encoding a hemol...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/1574-6968.12601
更新日期:2014-11-01 00:00:00
abstract::We investigated whether a subinhibitory concentration of clindamycin (Cli), corresponding to 1/2 the strain-specific minimum inhibitory concentration (MIC), could affect expression and stability of transcripts from genes coding for specific adhesins such as fibronectin binding proteins A (fnbA) and B (fnbB) as well as...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/j.femsle.2005.08.022
更新日期:2005-11-01 00:00:00
abstract::Proteins which are secreted or associated with the cell envelope of Mycobacterium tuberculosis may contain protective T-cell epitopes. Prior to this study, a recombinant clone bank of enzymatically active M. tuberculosis-alkaline phosphatase fusions, were screened for immunogenicity in a murine T-cell model. Five of t...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1999.tb13722.x
更新日期:1999-08-01 00:00:00
abstract::Weissella cibaria isolated from human saliva produces a soluble glucan that predominantly has alpha-1,6-glucosidic type linkages. Using degenerated primers that were selected based on the amino acid sequences of conserved regions from known glucansucrases, a single 2.7-kb fragment was isolated. In subsequent steps, a ...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2008.01460.x
更新日期:2009-03-01 00:00:00
abstract::We tested the possibility of nonconjugative lateral DNA transfer in a colony biofilm of mixed Escherichia coli strains. By simply coculturing a plasmid-free F(-) strain and another F(-) strain harboring a nonconjugative plasmid in a colony biofilm on antibiotic-free agar media, transformed cells were produced within 2...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2005.00072.x
更新日期:2006-02-01 00:00:00
abstract::Using natural rubber latex as the sole source of carbon and energy 50 rubber-degrading bacteria were isolated. Out of those 50 isolates, 33 were identified as Streptomyces species and 8 as Micromonospora species. Screening of 1220 bacteria obtained from different culture collections revealed 46 additional rubber-degra...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/s0378-1097(97)00072-4
更新日期:1997-05-15 00:00:00
abstract::Pseudomonas aeruginosa PA96 is a clinical isolate from Guangzhou, China, that is multiresistant to antibiotics. We previously described the 500-kb IncP-2 plasmid, pOZ176 that encodes many resistance genes including the IMP-9 carbapenemase. Whole-genome sequencing of PA96 enabled characterization of its genomic islands...
journal_title:FEMS microbiology letters
pub_type: 信件
doi:10.1111/1574-6968.12435
更新日期:2014-07-01 00:00:00
abstract::Acetone-butanol-ethanol (ABE) fermentation is a metabolic process of clostridia that produces bio-based solvents including butanol. It is enabled by an underlying metabolic reaction network and modulated by cellular gene regulation and environmental cues. Mathematical modeling has served as a valuable strategy to faci...
journal_title:FEMS microbiology letters
pub_type: 杂志文章,评审
doi:10.1093/femsle/fnw074
更新日期:2016-05-01 00:00:00
abstract::Xylose reductase catalyzes the NAD(P)H-dependent reduction of xylose to xylitol and is essential for growth on xylose by yeasts. To understand the nature of coenzyme binding to the Pichia stipitis xylose reductase, we investigated the role of the strictly conserved Lys270 in the putative IPKS coenzyme binding motif by...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1998.tb12848.x
更新日期:1998-02-01 00:00:00
abstract::Within the phylum Bacteroidetes, the gyrB gene, encoding for the B subunit of the DNA gyrase, has been used as a phylogenetic marker for several genera closely related to Flavobacterium. The phylogenies of the complete 16S rRNA gene and the gyrB gene were compared for 33 Antarctic Flavobacterium isolates and 23 type s...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2011.02326.x
更新日期:2011-08-01 00:00:00
abstract::Fifty-five geographically different strains of Fusarium poae were assayed for the presence of extrachromosomal nucleic acid elements. All strains were found to harbour double-stranded RNA (dsRNA) elements and encapsidated virus-like particles (VLP). There were great individual differences in dsRNA patterns of the vari...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1995.tb07790.x
更新日期:1995-09-15 00:00:00
abstract::The development of natural antibodies to Lactococcus lactis in three inbred strains of mice and the effect of inoculating L. lactis into these mice has been investigated. All animals developed detectable levels of natural (systemic and secretory) anti-lactococcal antibodies. Systemic anti-lactococcal antibodies were p...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1994.tb07041.x
更新日期:1994-07-15 00:00:00
abstract::A novel bacterial strain, PM2, capable of growing on methanol, was isolated in alkaline conditions from a soil inoculum. This bacterium was characterized at the physiological, biochemical and molecular level. Based on biochemical and molecular data strain PM2 was classified as a novel member of the group of fluorescen...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/S0378-1097(03)00692-X
更新日期:2003-10-24 00:00:00
abstract::Picrophilus torridus has been shown to degrade glucose via a nonphosphorylative Entner-Doudoroff (ED) pathway. Here we report the characterization of a key enzyme of this pathway, glycerate kinase (2-phosphoglycerate forming). The enzyme was purified 5,100-fold to homogeneity. The 95 kDa homodimeric protein catalyzed ...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2006.00264.x
更新日期:2006-06-01 00:00:00
abstract::The number and incidence of Collinsella aerofaciens in the human intestine are the highest among Gram-positive non-spore-forming bacilli. Identification of this species is very difficult and requires considerable time. A PCR-based identification system using C. aerofaciens-specific primers is described. Using this PCR...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2000.tb08931.x
更新日期:2000-02-01 00:00:00
abstract::Chromosomal DNA from 62 strains of 'Campylobacter upsaliensis' from blood and faecal specimens of patients in Australia, Belgium, South Africa and England, and seven dog isolates from England and Sweden were examined. DNA base compositions of 21 strains were 35 +/- 1 mol% G + C. Electrophoresis of HaeIII DNA digests g...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/0378-1097(90)90335-n
更新日期:1990-10-01 00:00:00
abstract::The existence in our laboratory of over 10000 Salmonella typhimurium LT2 cultures sealed in agar stab vials for 33-46 years offers an opportunity for evolutionary and mutational studies. In each of 77 vials examined, 10(3)-10(5) colony forming units per vial were recovered (less than 0.01% of the original population) ...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2001.tb10677.x
更新日期:2001-05-30 00:00:00
abstract::Periodic evolution of H(2)S during aerobic chemostat culture of Saccharomyces cerevisiae resulted in ultradian metabolic oscillation via periodic inhibition of respiratory activity. To understand the nature of periodic H(2)S evolution, we investigated whether oxidative stress is associated with H(2)S production. The c...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/S0378-1097(02)01198-9
更新日期:2003-02-14 00:00:00
abstract::The est gene encoding an esterase from Acinetobacter lwoffii RAG-1 was cloned into E. coli under the control of the PL promoter of the phage lambda. The N-terminal sequence of the first 20 amino acids of the heterologous expressed esterase corresponded to that obtained from the nucleotide sequence. Antibodies prepared...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1993.tb06462.x
更新日期:1993-09-15 00:00:00
abstract::beta-Glucosidase in Aspergillus nidulans was found to be both intracellular and extracellular. The intracellular beta-glucosidase was synthesized after the exhaustion of carbon source in the medium. The extracellular enzyme appeared with autolysis of the mycelium. Biosynthesis of beta-glucosidase was not induced by va...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1996.tb07969.x
更新日期:1996-01-01 00:00:00
abstract::Molecular microbial ecology studies are heavily reliant on 'Universal' 16S rRNA gene primers for elucidating microbial community structure and composition, and yet primer design and optimization is often overlooked. Primers that exhibit minor biases due to primer-template mismatches can substantially alter the pool of...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2011.02302.x
更新日期:2011-07-01 00:00:00
abstract::A different mutation (SMR1B) to SMR1-410 (ilv2), which determines resistance to sulfometuron methyl in Saccharomyces cerevisiae, was cloned by PCR. Sequence analysis indicated a C to T change at position 575 of the ILV2 coding sequence, which results in a proline transition to leucine at position 192. Similarly to SMR...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.1996.tb08100.x
更新日期:1996-04-01 00:00:00
abstract::PhoP was previously shown to be important for Yersinia pestis survival in macrophage and under macrophage-induced stresses. In this work, a phoP disruptant of Y. pestis 201 was generated using the Red cloning procedure. The transcription profile of the wild-type Y. pestis was then compared with that of the phoP mutant...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/j.femsle.2005.06.053
更新日期:2005-09-01 00:00:00
abstract::The cyanobacterium Synechocystis 6803 was tagged by chromosomal integration of the firefly luciferase gene, luc, resulting in the modified strain Synechocystis 6803-luc. The tagged cells were monitored in Baltic Sea microcosms both by detection of the luc gene by PCR amplification and by measurement of luc gene expres...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/0378-1097(95)00133-P
更新日期:1995-06-01 00:00:00
abstract::The gacA gene of the biocontrol strain Pseudomonas fluorescens CHA0 codes for a response regulator which, together with the sensor kinase GacS (=LemA), is required for the production of exoenzymes and secondary metabolites involved in biocontrol, including hydrogen cyanide (HCN). A gacA multicopy suppressor was isolat...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2000.tb09136.x
更新日期:2000-06-01 00:00:00
abstract::Aspergillus niger represents a promising host for the expression of recombinant proteins, but only a few expression systems are available for this organism. In this study, the inducible catalase promoter (PcatR) from A. niger was characterized. For this, constructs were developed and checked for the expression of the ...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2011.02454.x
更新日期:2012-02-01 00:00:00
abstract::Uncultured archaeal anaerobic methanotrophs (ANMEs) are known to operate the anaerobic oxidation of methane process, an important sink for the greenhouse gas methane in natural environments. In this study, we designed 16S rRNA gene-specific primers for each of the phylogenetic groups of ANMEs (ANME-1, Guaymas Basin hy...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2009.01648.x
更新日期:2009-08-01 00:00:00
abstract::Corynebacterium diphtheriae, generally considered an extracellular coloniser, was evaluated for its ability to enter and survive within HEp-2 monolayers by gentamicin protection assay. Intracellular viability of HC01 strain, isolated from endocarditis, was more expressive (2.59%) than observed in 241 (0.21%) and CDC-E...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2002.tb11379.x
更新日期:2002-09-24 00:00:00
abstract::We have performed amplified-fragment length polymorphism (AFLP) fingerprinting on a collection of Salmonella enterica subsp. enterica serovar typhimurium strains with a restriction endonuclease combination (BglII and MfeI) that has previously been used successfully for typing Campylobacter jejuni isolates with high re...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1111/j.1574-6968.2000.tb09200.x
更新日期:2000-08-01 00:00:00
abstract::Small colony variants (SCVs) of Pseudomonas aeruginosa NCTC 6750 (WT) were repeatedly isolated in an in vitro kinetic model after exposure to gentamicin (GM). There were minor differences biochemically and in phage and serotyping between the wild type (WT) strain and SCVs. Changes in outer membrane protein profiles we...
journal_title:FEMS microbiology letters
pub_type: 杂志文章
doi:10.1016/0378-1097(89)90165-1
更新日期:1989-10-01 00:00:00