Cloning, production, and functional expression of the bacteriocin sakacin A (SakA) and two SakA-derived chimeras in lactic acid bacteria (LAB) and the yeasts Pichia pastoris and Kluyveromyces lactis

2013 ◽  
Vol 40 (9) ◽  
pp. 977-993 ◽  
Author(s):  
Juan J. Jiménez ◽  
Juan Borrero ◽  
Dzung B. Diep ◽  
Loreto Gútiez ◽  
Ingolf F. Nes ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Pichia Pastoris ◽  
Lactic Acid Bacteria ◽  
Kluyveromyces Lactis ◽  
Functional Expression ◽  
Sakacin A

scholarly journals Functional expression of the lactate permease Jen1p of Saccharomyces cerevisiae in Pichia pastoris

2003 ◽  
Vol 376 (3) ◽  
pp. 781-787 ◽  
Author(s):  
Isabel SOARES-SILVA ◽  
Dorit SCHULLER ◽  
Raquel P. ANDRADE ◽  
Fátima BALTAZAR ◽  
Fernanda CÁSSIO ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Acetic Acid ◽  
Lactic Acid ◽  
Pichia Pastoris ◽  
Plasma Membranes ◽  
Membrane Vesicle ◽  
Dry Weight ◽  
Functional Expression ◽  
Acid Uptake ◽  
Methanol Induction

In Saccharomyces cerevisiae the activity for the lactate–proton symporter is dependent on JEN1 gene expression. Pichia pastoris was transformed with an integrative plasmid containing the JEN1 gene. After 24 h of methanol induction, Northern and Western blotting analyses indicated the expression of JEN1 in the transformants. Lactate permease activity was obtained in P. pastoris cells with a Vmax of 2.1 nmol·s−1·mg of dry weight−1. Reconstitution of the lactate permease activity was achieved by fusing plasma membranes of P. pastoris methanol-induced cells with Escherichia coli liposomes containing cytochrome c oxidase, as proton-motive force. These assays in reconstituted heterologous P. pastoris membrane vesicles demonstrate that S. cerevisiae Jen1p is a functional lactate transporter. Moreover, a S. cerevisiae strain deleted in the JEN1 gene was transformed with a centromeric plasmid containing JEN1 under the control of the glyceraldehyde-3-phosphate dehydrogenase constitutive promotor. Constitutive JEN1 expression and lactic acid uptake were observed in cells grown on either glucose and/or acetic acid. The highest Vmax (0.84 nmol·s−1·mg of dry weight−1) was obtained in acetic acid-grown cells. Thus overexpression of the S. cerevisiae JEN1 gene in both S. cerevisiae and P. pastoris cells resulted in increased activity of lactate transport when compared with the data previously reported in lactic acid-grown cells of native S. cerevisiae strains. Jen1p is the only S. cerevisiae secondary porter characterized so far by heterologous expression in P. pastoris at both the cell and the membrane-vesicle levels.

scholarly journals The characteristic of sheep cheese “Bryndza” from different regions of Slovakia based on microbiological quality

10.5219/1239 ◽  
2020 ◽  
Vol 14 ◽  
pp. 69-75 ◽  
Author(s):  
Miroslava Kačániová ◽  
Ľudmila Nagyová ◽  
Jana Štefániková ◽  
Soňa Felsöciová ◽  
Lucia Godočíková ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Kluyveromyces Lactis ◽  
Microbiological Quality ◽  
Aerobic Condition ◽  
Plate Count ◽  
Total Bacterial Count ◽  
Microscopic Fungi ◽  
Gram Positive ◽  
Gram Negative

The aim of our study was to describe microorganisms which occur in the traditional Slovak cheese „Bryndza“. There were a total of 60 cheese samples collected from ten different farms during May 2019. The microbiota studies included the total bacterial count, coliforms, enterococci, lactic acid bacteria, yeasts and microscopic fungi. The total bacterial counts were cultivated on plate count agar at 30 °C in aerobic conditions, lactic acid bacteria on MRS at 37 °C in anaerobic conditions, coliform on VRBL and VRBG at 37 °C in aerobic condition, yeasts and microscopic fungi on MEA at 25 °C under aerobic condition. Gram-positive, Gram-negative and yeasts isolates were identified with MALDI-TOF MS Biotyper. Totally, a number of 1175 isolates of G-, G+ and yeast were identified with score higher than 2 and moulds. Escherichia coli and Stenotrophomonas maltophilia were the most frequently identified species of Gram-negative and Leuconostoc mesenteroides ssp. mesenteroides and Lactococcus lactis ssp. lactis from Gram-positive bacteria. Yarrowia lipolitica and Kluyveromyces lactis were the most distributed yeasts. Lactic acid bacteria group was represented by Lactobacillus, Lactococcus, Leuconostoc and Pediococcus. The most abundant genera of lactic acid bacteria were Lactobacillus with 11 species. This study describes the indigenous microbiota of the traditional ewe's milk cheeses from Slovakia.

scholarly journals Cloning, Production, and Functional Expression of the Bacteriocin Enterocin A, Produced by Enterococcus faecium T136, by the Yeasts Pichia pastoris, Kluyveromyces lactis, Hansenula polymorpha, and Arxula adeninivorans

2012 ◽  
Vol 78 (16) ◽  
pp. 5956-5961 ◽  
Author(s):  
Juan Borrero ◽  
Gotthard Kunze ◽  
Juan J. Jiménez ◽  
Erik Böer ◽  
Loreto Gútiez ◽  
...  
Keyword(s):  
Pichia Pastoris ◽  
Enterococcus Faecium ◽  
Kluyveromyces Lactis ◽  
Hansenula Polymorpha ◽  
Functional Expression ◽  
Antimicrobial Activities ◽  
Arxula Adeninivorans ◽  
Content Type ◽  
Enterocin A

ABSTRACTThe bacteriocin enterocin A (EntA) produced byEnterococcus faeciumT136 has been successfully cloned and produced by the yeastsPichia pastorisX-33EA,Kluyveromyces lactisGG799EA,Hansenula polymorphaKL8-1EA, andArxula adeninivoransG1212EA. Moreover,P. pastorisX-33EA andK. lactisGG799EA produced EntA in larger amounts and with higher antimicrobial and specific antimicrobial activities than the EntA produced byE. faeciumT136.

scholarly journals Cloning and Heterologous Production of Hiracin JM79, a Sec-Dependent Bacteriocin Produced by Enterococcus hirae DCH5, in Lactic Acid Bacteria and Pichia pastoris

2008 ◽  
Vol 74 (8) ◽  
pp. 2471-2479 ◽  
Author(s):  
Jorge Sánchez ◽  
Juan Borrero ◽  
Beatriz Gómez-Sala ◽  
Antonio Basanta ◽  
Carmen Herranz ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Pichia Pastoris ◽  
Lactic Acid Bacteria ◽  
Expression Vector ◽  
Antimicrobial Activities ◽  
Biologically Active ◽  
Lactobacillus Sakei ◽  
Heterologous Production ◽  
Enterococcus Hirae ◽  
Gene Encoding

ABSTRACT Hiracin JM79 (HirJM79), a Sec-dependent bacteriocin produced by Enterococcus hirae DCH5, was cloned and produced in Lactococcus lactis, Lactobacillus sakei, Enterococcus faecium, Enterococcus faecalis, and Pichia pastoris. For heterologous production of HirJM79 in lactic acid bacteria (LAB), the HirJM79 structural gene (hirJM79), with or without the HirJM79 immunity gene (hiriJM79), was cloned into the plasmid pMG36c under the control of the constitutive promoter P32 and into the plasmid pNZ8048 under the control of the inducible PNisA promoter. For the production of HirJM79 in P. pastoris, the gene encoding the mature HirJM79 protein was cloned into the pPICZαA expression vector. The recombinant plasmids permitted the production of biologically active HirJM79 in the supernatants of L. lactis IL1403, L. lactis NZ9000, L. sakei Lb790, E. faecalis JH2-2, and P. pastoris X-33, the coproduction of HirJM79 and nisin A in L. lactis DPC5598, and the coproduction of HirJM79 and enterocin P in E. faecium L50/14-2. All recombinant LAB produced larger quantities of HirJM79 than E. hirae DCH5, although the antimicrobial activities of most transformants were lower than that predicted from their production of HirJM79. The synthesis, processing, and secretion of HirJM79 proceed efficiently in recombinant LAB strains and P. pastoris.

scholarly journals Sensory Improvement of a Pea Protein-Based Product Using Microbial Co-Cultures of Lactic Acid Bacteria and Yeasts

Foods ◽  
2020 ◽  
Vol 9 (3) ◽  
pp. 349 ◽  
Author(s):  
Cynthia El Youssef ◽  
Pascal Bonnarme ◽  
Sébastien Fraud ◽  
Anne-Claire Péron ◽  
Sandra Helinck ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Kluyveromyces Lactis ◽  
Starter Culture ◽  
Flavor Compounds ◽  
Protein Solution ◽  
Sensory Panel ◽  
Pea Protein ◽  
Off Flavors ◽  
Pea Proteins

Consumer demands for plant-based products have increased in recent years. However, their consumption is still limited due to the presence of off-flavor compounds, primarily beany and green notes, which are mainly associated with the presence of aldehydes, ketones, furans, and alcohols. To overcome this problem, fermentation is used as a lever to reduce off-flavors. A starter culture of lactic acid bacteria (LAB) was tested in a 4% pea protein solution with one of the following yeasts: Kluyveromyces lactis, Kluyveromyces marxianus, or Torulaspora delbrueckii. The fermented samples were evaluated by a sensory panel. Non-fermented and fermented matrices were analyzed by gas chromatography coupled with mass spectrometry to identify and quantify the volatile compounds. The sensory evaluation showed a significant reduction in the green/leguminous attributes of pea proteins and the generation of new descriptors in the presence of yeasts. Compared to the non-fermented matrix, fermentations with LAB or LAB and yeasts led to the degradation of many off-flavor compounds. Moreover, the presence of yeasts triggered the generation of esters. Thus, fermentation by a co-culture of LAB and yeasts can be used as a powerful tool for the improvement of the sensory perception of a pea protein-based product.

Synthesis of Mannose-Rich Exopolysaccharide by Rhodotorula glutinis 16P Co-Cultured with Yeast or Bacteria

2000 ◽  
Vol 55 (7-8) ◽  
pp. 540-545 ◽  
Author(s):  
Emilina D. Simova ◽  
Ginka I. Frengova ◽  
Dora M. Beshkova
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Cheese Whey ◽  
Kluyveromyces Lactis ◽  
Maximum Yield ◽  
Rhodotorula Glutinis ◽  
Lactobacillus Helveticus ◽  
Lactose Hydrolysis ◽  
Carbohydrate Composition ◽  
Structural Units

Abstract Exopolysaccharides from the lactose-negative yeast Rhodotorula glutinis 16P were synthesized by co-cultivation with the yeast Kluyveromyces lactis MP11 or with the homofermentative lactic acid bacteria Lactobacillus helveticus 9A in a cheese whey ultrafiltrate. Exopolysaccharides were produced by lactose hydrolysis, performed by two pathways: with β-galactosidase from Kluyveromyces lactis MP11 which assimilates glucose and galactose; with β-galactosidase and Lactobacillus helveticus 9A which uses lactic acid. By growing the two mixed cultures maximum yield was obtained as follows: 11.4 g/l and 15.8 g/l, respectively. Structural units of the carbohydrate composition of the two polymers are mannose (72.4-63.5%), glucose (2.0-15.9%), galactose (25.3-19.8%) and xylose (3.6-4.3%). Mannose dominated in the polysaccharide compositions.

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