scholarly journals Human ERCC5 cDNA-cosmid complementation for excision repair and bipartite amino acid domains conserved with RAD proteins of Saccharomyces cerevisiae and Schizosaccharomyces pombe.

1993 ◽  
Vol 13 (10) ◽  
pp. 6393-6402 ◽  
Author(s):  
M A MacInnes ◽  
J A Dickson ◽  
R R Hernandez ◽  
D Learmonth ◽  
G Y Lin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Excision Repair ◽  
Cdna Clones ◽  
Recombination Mechanism ◽  
Helix Loop Helix ◽  
Partial Length ◽  
Human Genes ◽  
Single Clone

Several human genes related to DNA excision repair (ER) have been isolated via ER cross-species complementation (ERCC) of UV-sensitive CHO cells. We have now isolated and characterized cDNAs for the human ERCC5 gene that complement CHO UV135 cells. The ERCC5 mRNA size is about 4.6 kb. Our available cDNA clones are partial length, and no single clone was active for UV135 complementation. When cDNAs were mixed pairwise with a cosmid clone containing an overlapping 5'-end segment of the ERCC5 gene, DNA transfer produced UV-resistant colonies with 60 to 95% correction of UV resistance relative to either a genomic ERCC5 DNA transformant or the CHO AA8 progenitor cells. cDNA-cosmid transformants regained intermediate levels (20 to 45%) of ER-dependent reactivation of a UV-damaged pSVCATgpt reporter plasmid. Our evidence strongly implicates an in situ recombination mechanism in cDNA-cosmid complementation for ER. The complete deduced amino acid sequence of ERCC5 was reconstructed from several cDNA clones encoding a predicted protein of 1,186 amino acids. The ERCC5 protein has extensive sequence similarities, in bipartite domains A and B, to products of RAD repair genes of two yeasts, Saccharomyces cerevisiae RAD2 and Schizosaccharomyces pombe rad13. Sequence, structural, and functional data taken together indicate that ERCC5 and its relatives are probable functional homologs. A second locus represented by S. cerevisiae YKL510 and S. pombe rad2 genes is structurally distinct from the ERCC5 locus but retains vestigial A and B domain similarities. Our analyses suggest that ERCC5 is a nuclear-localized protein with one or more highly conserved helix-loop-helix segments within domains A and B.

Human ERCC5 cDNA-cosmid complementation for excision repair and bipartite amino acid domains conserved with RAD proteins of Saccharomyces cerevisiae and Schizosaccharomyces pombe

1993 ◽  
Vol 13 (10) ◽  
pp. 6393-6402
Author(s):  
M A MacInnes ◽  
J A Dickson ◽  
R R Hernandez ◽  
D Learmonth ◽  
G Y Lin ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Excision Repair ◽  
Cdna Clones ◽  
Recombination Mechanism ◽  
Helix Loop Helix ◽  
Partial Length ◽  
Human Genes ◽  
Single Clone

Several human genes related to DNA excision repair (ER) have been isolated via ER cross-species complementation (ERCC) of UV-sensitive CHO cells. We have now isolated and characterized cDNAs for the human ERCC5 gene that complement CHO UV135 cells. The ERCC5 mRNA size is about 4.6 kb. Our available cDNA clones are partial length, and no single clone was active for UV135 complementation. When cDNAs were mixed pairwise with a cosmid clone containing an overlapping 5'-end segment of the ERCC5 gene, DNA transfer produced UV-resistant colonies with 60 to 95% correction of UV resistance relative to either a genomic ERCC5 DNA transformant or the CHO AA8 progenitor cells. cDNA-cosmid transformants regained intermediate levels (20 to 45%) of ER-dependent reactivation of a UV-damaged pSVCATgpt reporter plasmid. Our evidence strongly implicates an in situ recombination mechanism in cDNA-cosmid complementation for ER. The complete deduced amino acid sequence of ERCC5 was reconstructed from several cDNA clones encoding a predicted protein of 1,186 amino acids. The ERCC5 protein has extensive sequence similarities, in bipartite domains A and B, to products of RAD repair genes of two yeasts, Saccharomyces cerevisiae RAD2 and Schizosaccharomyces pombe rad13. Sequence, structural, and functional data taken together indicate that ERCC5 and its relatives are probable functional homologs. A second locus represented by S. cerevisiae YKL510 and S. pombe rad2 genes is structurally distinct from the ERCC5 locus but retains vestigial A and B domain similarities. Our analyses suggest that ERCC5 is a nuclear-localized protein with one or more highly conserved helix-loop-helix segments within domains A and B.

scholarly journals The Schizosaccharomyces pombe homolog of Saccharomyces cerevisiae HAP2 reveals selective and stringent conservation of the small essential core protein domain.

1991 ◽  
Vol 11 (2) ◽  
pp. 611-619 ◽  
Author(s):  
J T Olesen ◽  
J D Fikes ◽  
L Guarente
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Transcriptional Activation ◽  
Core Protein ◽  
Protein Domain ◽  
Yeast Saccharomyces Cerevisiae ◽  
Acid Protein ◽  
Ccaat Box

The fission yeast Schizosaccharomyces pombe is immensely diverged from budding yeast (Saccharomyces cerevisiae) on an evolutionary time scale. We have used a fission yeast library to clone a homolog of S. cerevisiae HAP2, which along with HAP3 and HAP4 forms a transcriptional activation complex that binds to the CCAAT box. The S. pombe homolog php2 (S. pombe HAP2) was obtained by functional complementation in an S. cerevisiae hap2 mutant and retains the ability to associate with HAP3 and HAP4. We have previously demonstrated that the HAP2 subunit of the CCAAT-binding transcriptional activation complex from S. cerevisiae contains a 65-amino-acid "essential core" structure that is divisible into subunit association and DNA recognition domains. Here we show that Php2 contains a 60-amino-acid block that is 82% identical to this core. The remainder of the 334-amino-acid protein is completely without homology to HAP2. The function of php2 in S. pombe was investigated by disrupting the gene. Strikingly, like HAP2 in S. cerevisiae, the S. pombe gene is specifically involved in mitochondrial function. This contrasts to the situation in mammals, in which the homologous CCAAT-binding complex is a global transcriptional activator.

scholarly journals Functional Expression and Characterization of Schizosaccharomyces pombe Avt3p as a Vacuolar Amino Acid Exporter in Saccharomyces cerevisiae

PLoS ONE ◽  
2015 ◽  
Vol 10 (6) ◽  
pp. e0130542 ◽  
Author(s):  
Soracom Chardwiriyapreecha ◽  
Kunio Manabe ◽  
Tomoko Iwaki ◽  
Miyuki Kawano-Kawada ◽  
Takayuki Sekito ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Functional Expression

The Schizosaccharomyces pombe homolog of Saccharomyces cerevisiae HAP2 reveals selective and stringent conservation of the small essential core protein domain

1991 ◽  
Vol 11 (2) ◽  
pp. 611-619
Author(s):  
J T Olesen ◽  
J D Fikes ◽  
L Guarente
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Fission Yeast ◽  
Transcriptional Activation ◽  
Core Protein ◽  
Protein Domain ◽  
Yeast Saccharomyces Cerevisiae ◽  
Acid Protein ◽  
Ccaat Box

The fission yeast Schizosaccharomyces pombe is immensely diverged from budding yeast (Saccharomyces cerevisiae) on an evolutionary time scale. We have used a fission yeast library to clone a homolog of S. cerevisiae HAP2, which along with HAP3 and HAP4 forms a transcriptional activation complex that binds to the CCAAT box. The S. pombe homolog php2 (S. pombe HAP2) was obtained by functional complementation in an S. cerevisiae hap2 mutant and retains the ability to associate with HAP3 and HAP4. We have previously demonstrated that the HAP2 subunit of the CCAAT-binding transcriptional activation complex from S. cerevisiae contains a 65-amino-acid "essential core" structure that is divisible into subunit association and DNA recognition domains. Here we show that Php2 contains a 60-amino-acid block that is 82% identical to this core. The remainder of the 334-amino-acid protein is completely without homology to HAP2. The function of php2 in S. pombe was investigated by disrupting the gene. Strikingly, like HAP2 in S. cerevisiae, the S. pombe gene is specifically involved in mitochondrial function. This contrasts to the situation in mammals, in which the homologous CCAAT-binding complex is a global transcriptional activator.

scholarly journals Molecular Characterization and Heterologous Expression of the Gene Encoding a Low-Molecular-Mass Endoglucanase from Trichoderma reesei QM9414

1998 ◽  
Vol 64 (2) ◽  
pp. 555-563 ◽  
Author(s):  
Hirofumi Okada ◽  
Kohji Tada ◽  
Tadashi Sekiya ◽  
Kengo Yokoyama ◽  
Akinori Takahashi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Schizosaccharomyces Pombe ◽  
Molecular Mass ◽  
Trichoderma Reesei ◽  
Active Form ◽  
Leader Peptide ◽  
Cdna Clones ◽  
Gene Transcript ◽  
E Coli ◽  
Molecular Masses

ABSTRACT We have isolated the genomic and cDNA clones encoding EG III (a low-molecular-mass endo-β-1,4-glucanase) gene fromTrichoderma reesei QM9414. The nucleotide sequence of the cDNA fragment was verified to contain a 702-bp open reading frame that encodes a 234-amino-acid propeptide. The deduced protein sequence has significant homologies with family H endo-β-1,4-glucanases. The 16-amino-acid N-terminal sequence was shown to function as a leader peptide for possible secretion. Northern blot analysis showed that the EG III gene transcript, with a length of about 700 bp, was expressed markedly by cellulose but not by glucose. The protein has been expressed as a mature form in Escherichia coli and as secreted forms in Saccharomyces cerevisiae and Schizosaccharomyces pombe under the control of tac, alcohol dehydrogenase (ADH1), and human cytomegalovirus promoters, respectively. The S. cerevisiae and Schizosaccharomyces pombe recombinant strains showed strong cellulolytic activities on agar plates containing carboxymethyl cellulose. The E. coli strain expressed small amounts of EG III in an active form and large amounts of EG III in an inactive form. The molecular masses of the recombinant EG IIIs were estimated to be 25, 28, and 29 kDa for E. coli, S. cerevisiae, and Schizosaccharomyces pombe, respectively, by immunoblot analysis following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Parts of the yeast recombinant EG IIIs decreased their molecular masses to 25 kDa after treatment with endoglycosidase H and α-mannosidase, suggesting that they are N glycosylated at least partly.

scholarly journals The rad18 gene of Schizosaccharomyces pombe defines a new subgroup of the SMC superfamily involved in DNA repair.

1995 ◽  
Vol 15 (12) ◽  
pp. 7067-7080 ◽  
Author(s):  
A R Lehmann ◽  
M Walicka ◽  
D J Griffiths ◽  
J M Murray ◽  
F Z Watts ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dna Repair ◽  
Schizosaccharomyces Pombe ◽  
Gene Deletion ◽  
Excision Repair ◽  
Repair Pathway ◽  
Repair Gene ◽  
Dna Repair Gene ◽  
Nucleotide Excision ◽  
Biochemical Analyses

The rad18 mutant of Schizosaccharomyces pombe is very sensitive to killing by both UV and gamma radiation. We have cloned and sequenced the rad18 gene and isolated and sequenced its homolog from Saccharomyces cerevisiae, designated RHC18. The predicted Rad18 protein has all the structural properties characteristic of the SMC family of proteins, suggesting a motor function--the first implicated in DNA repair. Gene deletion shows that both rad18 and RHC18 are essential for proliferation. Genetic and biochemical analyses suggest that the product of the rad18 gene acts in a DNA repair pathway for removal of UV-induced DNA damage that is distinct from classical nucleotide excision repair. This second repair pathway involves the products of the rhp51 gene (the homolog of the RAD51 gene of S. cerevisiae) and the rad2 gene.

The rad16 gene of Schizosaccharomyces pombe: a homolog of the RAD1 gene of Saccharomyces cerevisiae

1994 ◽  
Vol 14 (3) ◽  
pp. 2029-2040
Author(s):  
A M Carr ◽  
H Schmidt ◽  
S Kirchhoff ◽  
W J Muriel ◽  
K S Sheldrick ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Schizosaccharomyces Pombe ◽  
Mating Type ◽  
Excision Repair ◽  
Radiation Sensitivity ◽  
Reading Frame ◽  
Uv Sensitivity ◽  
Protein Protein Interaction ◽  
Mating Type Switching

The rad10, rad16, rad20, and swi9 mutants of the fission yeast Schizosaccharomyces pombe, isolated by their radiation sensitivity or abnormal mating-type switching, have been shown previously to be allelic. We have cloned DNA correcting the UV sensitivity or mating-type switching phenotype of these mutants and shown that the correcting DNA is encompassed in a single open reading frame. The gene, which we will refer to as rad16, is approximately 3 kb in length, contains seven introns, and encodes a protein of 892 amino acids. It is not essential for viability of S. pombe. The predicted protein is the homolog of the Saccharomyces cerevisiae RAD1 protein, which is involved in an early step in excision-repair of UV damage from DNA. The approximately 30% sequence identity between the predicted proteins from the two yeasts is distributed throughout the protein. Two-hybrid experiments indicate a strong protein-protein interaction between the products of the rad16 and swi10 genes of S. pombe, which mirrors that reported for RAD1 and RAD10 in S. cerevisiae. We have identified the mutations in the four alleles of rad16. They mapped to the N-terminal (rad10), central (rad20), and C-terminal (rad16 and swi9) regions. The rad10 and rad20 mutations are in the splice donor sequences of introns 2 and 4, respectively. The plasmid correcting the UV sensitivity of the rad20 mutation was missing the sequence corresponding to the 335 N-terminal amino acids of the predicted protein. Neither smaller nor larger truncations were, however, able to correct its UV sensitivity.

scholarly journals The rad16 gene of Schizosaccharomyces pombe: a homolog of the RAD1 gene of Saccharomyces cerevisiae.

1994 ◽  
Vol 14 (3) ◽  
pp. 2029-2040 ◽  
Author(s):  
A M Carr ◽  
H Schmidt ◽  
S Kirchhoff ◽  
W J Muriel ◽  
K S Sheldrick ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Schizosaccharomyces Pombe ◽  
Mating Type ◽  
Excision Repair ◽  
Radiation Sensitivity ◽  
Reading Frame ◽  
Uv Sensitivity ◽  
Protein Protein Interaction ◽  
Mating Type Switching

The rad10, rad16, rad20, and swi9 mutants of the fission yeast Schizosaccharomyces pombe, isolated by their radiation sensitivity or abnormal mating-type switching, have been shown previously to be allelic. We have cloned DNA correcting the UV sensitivity or mating-type switching phenotype of these mutants and shown that the correcting DNA is encompassed in a single open reading frame. The gene, which we will refer to as rad16, is approximately 3 kb in length, contains seven introns, and encodes a protein of 892 amino acids. It is not essential for viability of S. pombe. The predicted protein is the homolog of the Saccharomyces cerevisiae RAD1 protein, which is involved in an early step in excision-repair of UV damage from DNA. The approximately 30% sequence identity between the predicted proteins from the two yeasts is distributed throughout the protein. Two-hybrid experiments indicate a strong protein-protein interaction between the products of the rad16 and swi10 genes of S. pombe, which mirrors that reported for RAD1 and RAD10 in S. cerevisiae. We have identified the mutations in the four alleles of rad16. They mapped to the N-terminal (rad10), central (rad20), and C-terminal (rad16 and swi9) regions. The rad10 and rad20 mutations are in the splice donor sequences of introns 2 and 4, respectively. The plasmid correcting the UV sensitivity of the rad20 mutation was missing the sequence corresponding to the 335 N-terminal amino acids of the predicted protein. Neither smaller nor larger truncations were, however, able to correct its UV sensitivity.

Biological Processing of Pea Grain and Secondary Starch Raw Materials to Produce Food and Feed Protein Concentrates

2020 ◽  
Vol 36 (4) ◽  
pp. 49-58
Author(s):  
V.V. Kolpakova ◽  
R.V. Ulanova ◽  
L.V. Chumikina ◽  
V.V. Bessonov
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acid ◽  
Amino Acid Composition ◽  
Acid Composition ◽  
Mass Fraction ◽  
Geotrichum Candidum ◽  
Protein Concentrate ◽  
Protein Concentrates ◽  
Pea Flour ◽  
Nitrogenous Substances

The goal of the study was to develop a biotechnological process for the production of protein concentrates via bioconversion of pea flour and whey, a secondary product of starch manufacture. Standard and special methods were used to analyze the chemical and biochemical composition of protein concentrates (amino acid, carbohydrate, and fractional) of flour, whey and protein concentrates. It was established that pea flour contains 52.28-57.05% water-soluble nitrogenous substances, 23.04-25.50% salt-soluble, 2.94-4.69% alcohol-soluble compounds, 0-0.61% of soluble glutenine, 6.67-10.40% alkali-soluble glutenine and 5.96-10.86% insoluble sclerotic substances. A mathematical model and optimal parameters of the enzymatic extraction of pea protein with a yield of 65-70% were developed. Ultrasonic exposure increased the yield of nitrogenous substances by 23.16 ± 0.69%, compared with the control without ultrasound. The protein concentrate had a mass fraction of nitrogenous substances of 72.48 ± 0.41% (Nx6.25) and a complete amino acid composition. The microbial conversion by the Saccharomyces cerevisiae 121 and Geotrichum candidum 977 cultures of starch whey which remained after protein precipitation allowed us to obtain feed concentrates from biomass and culture liquid with a protein mass fraction of 61.68-70.48% (Nx6.25). Protein concentrates positively affected the vital signs of rats and their excretory products. A technological scheme was developed to test the complex pea grain and starch whey processing under pilot conditions. pea, protein concentrate, extracts, whey, bioconversion, Geotrichum candidum, Saccharomyces cerevisiae, chemical composition, amino acid composition

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