From DNA-DNA hybridization and guanine-plus-cytosine (G+C) contents of DNA in 46 strains of yeast-lysing bacteria isolated in Brazil, 15, 28 and 2 strains formed clusters with Japanese isolates YLM-32, YLM-115 and Rarobacter faecitabidus YLM-1T, respectively. The YLM-115 group had lysine as diamino acid in the cell wall peptidoglycan, and they remained unclassified. Rarobacter incanus, sp. nov., was proposed for the YLM-32 group strains. Their G+C contents of DNA were 64.6-65.5 mol%, which were slightly lower than those of R. faecitabidus. The amino acid composition of cell wall peptidoglycan of R. incanus was D-Ala, L-Ala, D-Glu and L-Orn (1:1:2:1), which closely resembled R. faecitabidus, although the latter contained a serine residue which the former did not. Other phenotypic and chemotaxonomic characteristics were almost the same as R. faecitabidus, but R. incanus formed a separate and distinct cluster from R. faecitabidus following DNA-DNA hybridization. The type strain is YLM-32 (JCM 6350).
Attempts were made to analyze the substances involved in the adhesion of cells of F. succinogenes S85 to cellulose. Firstly, the effect of the treatment of the cells with enzymes on their adhering ability to cellulose was examined. Treatment of cells with proteolytic enzymes such as proteinase K, thermolysin, protease Type IV and trypsin, and with lipase reduced markedly their adhering ability to cellulose, whereas their adhering ability to cellulose was not affected by treatment with acid phosphatase, DNase 1, RNase A and RNase T1. These results show that some proteins on the cell surface are involved in the adhesion of cells to cellulose. Next, attempts were made to detect some of the proteins capable of adhering to cellulose in cell lysate of the bacterium. After Avicel cellulose was incubated in the cell lysate of the bacterium at 37°C, cellulose was washed several times with buffer containing Softes 12 as a detergent to remove the cell debris and proteins incapable of adhering to cellulose. Proteins strongly adhering to cellulose after washing were eluted with buffers containing 1% carboxymethylcellulose (CMC) or 10% cellobiose. When the proteins eluted with CMC were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), one major band and many minor bands were detected in the gel. When the proteins eluted with cellobiose were analyzed by SDS-PAGE, one major band, one minor band and two faint bands were detected in the gel. The major protein in both eluents using CMC or cellobiose, exhibited the same molecular weight of approximately 120 kilodaltons (kDa). These CBPs in cells of the bacterium disappeared after treatment with proteinase K, whereas these CBPs in cells remained after treatment with lipase.
Strong expression system of yeast dominant selection marker gene was constructed for transformation of industrial yeasts. Bacterial aminoglycoside phosphotransferase II (APT2) gene from Escherichia coli transposon Tn903 was inserted between a yeast alcohol dehydrogenase I (ADH1) gene promoter and cytochrome c1 (CYC1) gene terminator. This ADH1-APT2 gene was strongly expressed in the transformed yeasts even in the case of integration type plasmid. Using this strong expressive dominant selection marker gene, five strains in nine industrial yeasts which were tested could not be transformed by lithium acetate method, but these industrial yeasts could be transformed by electroporation.
Estimates of the nuclear DNA (nDNA) content in the yeast cells of 43 strains of Leucosporidium scottii, Rhodosporidium toruloides, and related yeast taxa were analyzed by fluorescence microscope photometry. Quantitative differences of nDNA contents occur among 18 strains of L. scottii. Four groups of strains with similar nDNA content were recognizable in L. scottii. One group contained only one strain (IFO 9474) which was characterized by the Q-7 system. The respective strains of the remaining three groups had either Q-9 or Q-10 as the major ubiquinone system, except CBS 8188 having both of these. Some mating strains of L. scottii had almost twice the amount of nDNA as the presumed haploid value. In contrast, both mating type A and a strains of R. toruloides had nearly identical average nDNA contents. Our results suggest the presence of aneuploidy among yeast cells of L. scottii.
To establish the optimal conditions for mutagenesis in the astaxanthin-producing yeast Phaffia rhodozyma when different mutagens are used, dose-response curves for UV radiation, ethyl methanosulfonate and N-methyl-N′-nitro-N-nitrosoguanidine were constructed. An enrichment scheme using the polyene antibiotic nystatin for the isolation of auxotrophic mutants was designed. This treatment resulted in auxotrophic isolates at a frequency of 1×10-3. Uracil-requiring mutants were obtained by 5-fluoro-orotic acids selection at a frequency of 1.67×10-7.