Regulations of RLM1 gene affect the anti-autolytic ability of lager yeast / 生物工程学报

The autolysis of brewer's yeast seriously affects the quality of beer and the quality of yeast is considered as one of the key factors in beer brewing. Previous studies on brewer's yeast autolysis showed that RLM1 gene, an important transcription factor in cell integrity pathway, is closely related to the autolysis of yeast. In this study, RLM1 was knocked out and overexpressed in a haploid brewer's yeast. RLM1 disruption resulted in poor anti-autolysis performance of yeast, whereas overexpression of RLM1 contributed to the anti-autolytic ability of yeast. In addition, RLM1 gene knockout affected the osmotic stress resistance, cell wall damage resistance, nitrogen starvation resistance and temperature tolerance of yeast strain. The transcriptional level of GAS1 involved in cell wall assembly and DNA damage response was regulated along with the expression of RLM1, whereas other genes in CWI pathway did not show apparent regularity. RLM1 might mainly affect the expression of GAS1 so as to improve the stress resistance of lager yeast in harsh environment. The result from this study help further understand the mechanism of yeast autolysis and lay a foundation for breeding brewer's yeast strain with better anti-autolytic ability.

Biotransformation of bromhexine by Cunninghamella elegans, C. echinulata and C. blakesleeana

Abstract Fungi is a well-known model used to study drug metabolism and its production in in vitro condition. We aim to screen the most efficient strain of Cunninghamella sp. among C. elegans, C. echinulata and C. blakesleeana for bromhexine metabolites production. We characterized the metabolites produced using various analytical tools and compared them with mammalian metabolites in Rat liver microsomes (RLM). The metabolites were collected by two-stage fermentation of bromhexine with different strains of Cunninghamella sp. followed by extraction. Analysis was done by thin layer chromatography, high performance thin layer chromatography, Fourier transform infrared spectroscopy, high performance liquid chromatography and Liquid chromatography–mass spectrometry. The role of Cytochrome P3A4 (CYP3A4) enzymes in bromhexine metabolism was studied. Fungal incubates were spiked with reference standard – clarithromycin to confirm the role of CYP3A4 enzyme in bromhexine metabolism. Three metabolites appeared at 4.7, 5.5 and 6.4 min retention time in HPLC. Metabolites produced by C. elegans and RLM were concluded to be similar based on their retention time, peak area and peak response of 30.05%, 21.06%, 1.34%, and 47.66% of three metabolites and bromhexine in HPLC. The role of CYP3A4 enzyme in metabolism of bromhexine and the presence of these enzymes in Cunninghamella species was confirmed due to absence of peaks at 4.7, 5.4 and 6.7 min when RLM were incubated with a CYP3A4 enzyme inhibitor – clarithromycin.

The cloning and expression of human interleukin-17 receptor-like molecule / 西安交通大学学报(医学版)

Objective To generate an human interleukin-17 receptor-like molecule (IL-17RLM) recombinant plasmid with 6?myc tag and detect its specific expression in eukaryotic cells. Methods Design two specific primers(including the enzyme sites of EcoRⅠand XhoⅠ), reextract hIL-17RLM-L DNA fragment after PCR and insert it into the 6?myc tagged pcDNA3.0 vector, then detect its expression by Western blot after transfecting COS7 cells. Results The 6?myc tagged recombinant plasmid pcDNA3.0- 6?myc /hIL-17RLM-L was generated successfully and its expression can be detected by Western blot in eukaryotic cells. Conclusion The eukaryotic expressing plasmid pcDNA3.0-6?myc /hIL-17RLM-L was generated successfully and its specific expression was realized, which may provide the basis for further research of its biological function.