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S Hohmann

Publications and source records attributed to S Hohmann.

62 records · Page 4Linked to original sources
PubMed1989

A deletion of the PDC1 gene for pyruvate decarboxylase of yeast causes a different phenotype than previously isolated point mutations.

We deleted most of the pyruvate decarboxylase structural gene PDC1 from the genome of Saccharomyces cerevisiae. Surprisingly, mutants carrying this deletion allele showed a completely different phenotype than previously described point mutations. They were able to ferment glucose and their specific pyruvate decarboxylase activity was only reduced to 45% of the wild type level. Northern blot analysis revealed that a sequence in the yeast genome homologous to PDC1 and formerly designated as a possible pseudogene is expressed and may code for a different but closely related pyruvate decarboxylase. The products of the two PDC genes seem to form hybrid oligomers, however both homooligomers have enzyme activity. Thus, the product of the PDC1 gene is not absolutely necessary for glucose fermentation in yeast.

Blotting, Southern↗
PubMed1989

Comparison of the nucleotide sequences of a yeast gene family. I. Distribution and spectrum of spontaneous base substitutions.

The nucleotide sequences of closely related members of a gene family can be used to investigate spontaneous mutations. Here we analyse the sequences of different yeast invertase genes which are more than 93% identical in the coding region and share some very similar, but not identical sequences in the noncoding flanking regions. Since all except one of the invertase genes are active, most of the base substitutions are silent. Within the coding region the base substitutions are unevenly distributed, indicating that parts of the genes were homogenized, probably via gene conversion. Transitions occurred more frequently than transversions in both, coding and noncoding regions. In the coding region pyrimidine transitions were the most abundant event due to silent changes mainly in the third codon position. In the noncoding region pyrimidine and purine transitions were found at equal frequencies. Transversions inverting base pairs (A-T and G-C) outnumber transversions changing base pairs (A-C and G-T). While the spectrum of mutations in the coding region is influenced by selective pressure to maintain the amino acid sequence, the spectrum in the noncoding region may be much less affected by selective pressure.

Base Sequence↗
PubMed1989

Comparison of the nucleotide sequences of a yeast gene family. II. Analysis of spontaneous deletions and insertions.

We compared the nucleotide sequences of 3 yeast invertase genes in regions where the homology is better than 90%. In the noncoding region 40 gaps of 1-61 bases were found. This is about half as much as the nucleotide substitutions in the same sequences. We grouped the gaps into 5 categories by their length and the characteristics of their sequences. Group I gaps are about 20 nucleotides long and are flanked by repeated sequence of 6 bases which may trigger the deletion of one of the repeats and the sequence between the repeats. Group II gaps are characterized by a small repeated sequence which is missing in one of the invertase genes. Gaps which occur in sequences exclusively made up of one of the 4 bases are summarized in group III. The 4 gaps in group IV do not show any of these sequence characteristics and they are all just one base long. A 61 nucleotide sequence found in only one of the invertase genes seems to be of complex origin. We conclude that small repeated sequences or monotonous sequences are prone to deletion or insertion mutations.

Base Sequence↗
PubMed1988

Structural analysis of the 5' regions of yeast SUC genes revealed analogous palindromes in SUC, MAL and GAL.

In the yeast Saccharomyces cerevisiae six unlinked structural genes for invertase, the SUC genes, are known. We sequenced about 800 bp of the 5' non-coding region and the first 220 bp of the coding region of the genes SUC1, SUC3, SUC4 and SUC5 and compared them with the previously sequenced genes SUC2 and SUC7 (Sarokin and Carlson 1985a). All are highly homologous within the coding region but in the non-coding region SUC1 shows some differences and SUC2 is more highly diverged. Two different kinds of TATA boxes were identified: the more strongly expressed genes SUC1, 2 and 4 have the sequence TATAAA and the more weakly expressed genes SUC3, 5 and 7 have TACAAA. Though the SUC1 sequence is in general more homologous to the other SUC genes, the region between -140 and +100 of SUC1 is nearly identical to SUC2. This could be due to a gene conversion between SUC1 and the silent suc2 degrees allele which occurs in the strains carrying SUC1. Within the upstream regions of all the SUC genes three regions with palindromic sequences analogous to stem and loop structures were identified. Comparable structures could be detected in similar positions in the upstream sequences of the divergently transcribed yeast gene pairs MAL6S-MAL6T and GAL1-GAL10. Implications for the importance of these structures in the regulation and initiation of transcription are discussed.

Amino Acid Sequence↗
PubMed1987

A region in the yeast genome which favours multiple integration of DNA via homologous recombination.

Integrative transformation of yeast with gapped DNA fragments results in single or multiple integration into the yeast genome via homologous recombination. A sequence of yeast DNA was found which favours multiple integration even when the strategy of gene replacement is used. This strategy by which the transformed DNA fragment replaces its chromosomal homologue rather than simply integrating into the genome usually occurs as a single exchange event. The described region is unique and lies near a telomere about 5 kb proximal to the SUC4 locus on chromosome XIII. DNA from this region was used as a vehicle for the integration of different SUC genes coding for invertase. Most of the sucrose fermenting transformants isolated carried between two and seven copies of the SUC genes. These transformants overproduced invertase even though there was no selective pressure for high invertase activity in these experiments. I conclude that this region is highly recombinogenic and favours multiple integration of DNA fragments. This region could be used for stable multiple integration of heterologous genes into the yeast genome for over-production of the respective gene product.

Cloning, Molecular↗
PubMed1986

Cloning and expression on a multicopy vector of five invertase genes of Saccharomyces cerevisiae.

Six unlinked loci for invertase structural genes are known in the yeast Saccharomyces cerevisiae: SUC1-SUC5 and SUC7. These genes are similar in structure and expression but not identical. Different yeast strains possess none, one or several of these genes. We have isolated the genes SUC1-SUC5, subcloned them into the multicopy vector YEp24 and compared the expression of the five SUC genes in one recipient strain. SUC2 was isolated by transformation of a suc0 strain with a gene pool and complementation to sucrose fermentation. SUC4 was cloned from a minipool of chromosomal fragments which were shown to contain SUC4 by Southern hybridization. SUC1, SUC3 and SUC5 were isolated using the method of plasmid eviction. A plasmid containing regions flanking SUC4 was integrated next to these SUC genes. The plasmid together with the SUC genes were then cut out of the chromosome using an appropriate restriction endonuclease. The length of chromosomal DNA fragments containing the different SUC genes were 4.8 kb for SUC1, 5.2 kb for SUC2, 4.8 kb for SUC3, 12.8 kb for SUC4 and 17.2 kb for SUC5. Fragments containing the complete SUC genes and the sequences controlling their expression were subcloned into YEp24 and transformed into a strain without any active invertase gene. Invertase activity of transformants was measured after growth repressing (8% glucose) and derepressing (2% raffinose) conditions. As expected from results with strains carrying the individual SUC genes in a chromosomal location, the SUC genes were expressed to a different extent.

Cloning, Molecular↗