Search academic texts

Information × Registration Number 0220U104395, 0120U103573 , R & D reports Title Identification of genes involved in autophagic degradation of soluble proteins in yeast and human cells popup.stage_title Head Sybirnyi Andrii A., Registration Date 16-12-2020 Organization Institute of Cell Biology popup.description2 A selection system has been developed for identification of the strains of yeast Komagatella phaffii with deletion of the genes associated with autophagic degradation of cytosolic proteins. For this purpose, the LAC4 gene of Kluyveromyces lactis encoding β-galactosidase [Ishchuk et al., 2008], fused to the fluorescent label GFP, was expressed under the control of a methanol-induced promoter of the formaldehyde dehydrogenase gene in strains K. phaffii GS200 (arg4 his4) and SMD1163 (his4 pep4 prb1), in which the process of autophagy is disrupted. The obtained recombinant strains produced β-galactosidase during growth on medium with methanol, and after transfer of cells to medium with glucose, degradation of this cytosolic protein occurred. Because derivatives of strain SMD1163 showed a slower decrease in β-galactosidase activity than derivatives of strain GS200, it can be argued that the degradation of recombinant protein occurs in part by autophagy. Based on the engineered strain GS200 / LAC4, insertion mutants were obtained in which the process of β-galactosidase degradation was prolonged after transfer to glucose medium compared to the original strain. In one of the obtained mutants, the integration site of the insertion cassette was identified - the integration occurred in the region of the open reading frame of the gene encoding β-1,6-N-acetylglucosaminyltransferase. Product Description popup.authors Zazulya Anastasiya Zinoviivna Semkiv Marta Vitaliivna popup.nrat_date 2020-12-16 Close