Institute of Biochemistry and Biophysics, Polish Academy of Sciences, Pawińskiego 5a, 02-106 Warsaw, Poland.
Biol Chem. 2011 Apr;392(6):517-27. doi: 10.1515/BC.2011.058. Epub 2011 Apr 27.
In Trichoderma reesei, dolichyl phosphate mannose (dpm) synthase, a key enzyme in the O-glycosylation process, requires three proteins for full activity. In this study, the dpm2 and dpm3 genes coding for the DPMII and DPMIII subunits of T. reesei DPM synthase were cloned and functionally analyzed after expression in the Saccharomyces cerevisiae dpm1Δ [genotype (BY4743; his3Δ1; /leu2Δ0; lys2Δ0; /ura3Δ0; YPR183w::kanMX4] mutant. It was found that apart from the catalytic subunit DPMI, the DPMIII subunit is also essential to form an active DPM synthase in yeast. Additional expression of the DPMII protein, considered to be a regulatory subunit of DPM synthase, decreased the enzymatic activity. We also characterized S. cerevisiae strains expressing the dpm1, 2, 3 or dpm1, 3 genes and analyzed the consequences of dpm expression on protein O-glycosylation in vivo and on the cell wall composition.
在里氏木霉中，磷酸多萜醇甘露糖（dpm）合酶是 O-糖基化过程中的关键酶，需要三种蛋白质才能充分发挥活性。本研究在酿酒酵母 dpm1Δ [基因型（BY4743；his3Δ1；/ leu2Δ0； lys2Δ0；/ ura3Δ0；YPR183w::kanMX4]突变体中表达后，对编码里氏木霉 DPM 合酶 DPMII 和 DPMIII 亚基的 dpm2 和 dpm3 基因进行了克隆和功能分析。结果发现，除了催化亚基 DPMI 外，DPMIII 亚基对于在酵母中形成有活性的 DPM 合酶也是必需的。额外表达被认为是 DPM 合酶调节亚基的 DPMII 蛋白会降低酶活性。我们还对表达 dpm1、2、3 或 dpm1、3 基因的酿酒酵母菌株进行了表征，并分析了 dpm 表达对体内蛋白 O-糖基化和细胞壁组成的影响。
