Lipase localization in Rhizopus oryzae cells immobilized within biomass support particles for use as whole-cell biocatalysts in biodiesel-fuel production
Lipase localization in Rhizopus oryzae cells immobilized within biomass support particles for use as whole-cell biocatalysts in biodiesel-fuel production
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DOI：
10.1263/jbb.101.328
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发表时间：
2006-04-01
影响因子：
2.8
通讯作者：
Fukuda, Hideki
中科院分区：
文献类型：
--
作者：
Hama, Shinji;Tamalampudi, Sriappareddy;Fukuda, Hideki
为了鉴定负责真菌全细胞生物催化剂的甲醇分解活性的脂肪酶，确定了米根霉细胞的脂肪酶定位。 Western blot分析表明米根霉细胞产生两种不同分子量的脂肪酶，分别为34和31 kDa；前者（ROL34）与细胞壁结合，而后者（ROL31）主要与细胞膜结合。研究发现，网状聚氨酯泡沫生物质支撑颗粒内的细胞固定强烈抑制膜结合脂肪酶分泌到培养基中。对 ROL34 和 ROL31 之间关系的研究表明，ROL31 源自 ROL34 N 末端 28 个氨基酸残基的裂解。向培养基中添加橄榄油导致细胞内脂肪酶的保留量增加。通过菌丝细胞的免疫荧光标记进一步证实了这种现象。当用各种底物相关化合物（例如橄榄油和油酸）培养细胞时，细胞内甲醇分解活性与膜结合脂肪酶的相对量密切相关，这表明定位于膜的ROL31在米根霉细胞的甲醇分解活性中起着至关重要的作用。
To identify the lipase responsible for the methanolysis activity of fungus whole-cell biocatalysts, the lipase localization of Rhizopus oryzae cells was determined. Western blot analysis showed that R. oryzae cells produce two types of lipase with different molecular masses of 34 and 31 kDa; the former (ROL34) was bound to the cell wall, whereas the latter (ROL31) was mainly bound to the cell membrane. It was found that cell immobilization within reticulated polyurethane foam biomass support particles strongly inhibits the secretion of membrane-bound lipase into the culture medium. An investigation of the relationship between ROL34 and ROL31 suggested that ROL31 originates from the cleavage of a 28-amino-acid residue at the N-terminus of ROL34. The addition of olive oil to the culture medium led to the retention of increased amounts of lipase within the cell. This phenomenon was further confirmed by an immunofluorescence labeling of hyphal cells. When cells were cultivated with various substrate-related compounds, such as olive oil and oleic acid, the intracellular methanolysis activity strongly correlated with the relative amounts of the membrane-bound lipase, which suggests that ROL31 localized in the membrane plays a crucial role in the methanolysis activity of R. oryzae cells.