title: High-yield novel leech hyaluronidase to expedite the preparation of specific hyaluronan oligomers.
abstract: Hyaluronidases (HAases), particularly leech HAases, have attracted intense attention due to their broad applications in medical treatments and great potential for the enzymatic production of hyaluronan oligosaccharides. However, little is known about this third interesting family of HAases. Here, we applied the random amplification of cDNA ends polymerase chain reaction (RACE-PCR) approach to identify the first leech HAase-encoding gene. By combining protein engineering and high-density culture, we achieved high-level production (8.42 × 10(5) U ml(-1)) in the yeast Pichia pastoris secretory expression system. Compared with the commercial bovine testicular HAase, the recombinant leech HAase exhibited superior enzymatic properties. Furthermore, analysis of the hydrolytic process suggested that this novel enzyme adopts a nonprocessive endolytic mode, yielding a narrow-spectrum of specific HA oligosaccharides with different incubation times. Large-scale production of this novel leech HAase will not only greatly promote medical applications but also facilitate the enzymatic production of specific HA oligosaccharides.
authors.affiliation: 1] Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, P. R. China [2] The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, P. R. China [3] School of Biotechnology, Jiangnan University, Wuxi 214122, P. R. China.
authors.forename: Peng
authors.initials: P
authors.lastname: Jin
authors.affiliation: 1] Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, P. R. China [2] The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, P. R. China [3] School of Biotechnology, Jiangnan University, Wuxi 214122, P. R. China [4] Synergetic Innovation Center of Modern Industrial Fermentation, Wuxi 214122, P. R. China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Kang
authors.affiliation: 1] Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, P. R. China [2] The Key Laboratory of Industrial Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, P. R. China [3] School of Biotechnology, Jiangnan University, Wuxi 214122, P. R. China.
authors.forename: Na
authors.initials: N
authors.lastname: Zhang
authors.affiliation: 1] Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, P. R. China [2] School of Biotechnology, Jiangnan University, Wuxi 214122, P. R. China [3] The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, P. R. China [4] Synergetic Innovation Center of Modern Industrial Fermentation, Wuxi 214122, P. R. China.
authors.forename: Guocheng
authors.initials: G
authors.lastname: Du
authors.affiliation: 1] Synergetic Innovation Center of Food Safety and Nutrition, Jiangnan University, Wuxi 214122, P. R. China [2] School of Biotechnology, Jiangnan University, Wuxi 214122, P. R. China [3] National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, P. R. China [4] Synergetic Innovation Center of Modern Industrial Fermentation, Wuxi 214122, P. R. China.
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
mesh_terms: D000818:Animals; D002417:Cattle; D018076:DNA, Complementary; D006820:Hyaluronic Acid / Q000096:biosynthesis* / Q000737:chemistry; D006821:Hyaluronoglucosaminidase / Q000235:genetics / Q000378:metabolism*; D006868:Hydrolysis; D007865:Leeches / Q000201:enzymology*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis* / Q000737:chemistry; D010843:Pichia / Q000235:genetics
chemical_list: D018076:DNA, Complementary; D009844:Oligosaccharides; D006820:Hyaluronic Acid; D006821:Hyaluronoglucosaminidase