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Title 

Characterization of recombinant bovine sperm hyaluronidase and identification of an important Asn-X-Ser/Thr motif for its activity

Authors 

C ParkYoung-Hyun KimSang-Rae LeeS ParkY JungYoungjeon LeeJi-Su KimT EomJ S KimD M LeeBong-Seok SongBo Woong SimSun-Uk KimKyu Tae ChangE Kim

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

2018

Citation 

Journal of Microbiology and Biotechnology

Keywords 

Cumulus oocyte complexFertilizationHyaluronic acidHyaluronidase

Abstract 

Hyaluronidases are a family of enzymes that catalyse the breakdown of hyaluronic acid, which is abundant in the extracellular matrix and cumulus oocyte complex. To investigate the activity of recombinant bovine sperm hyaluronidase 1 (SPAM1) and determine the effect of the Asn-X-Ser/Thr motif on its activity, the bovine SPAM1 open reading frame was cloned into the mammalian expression vector pCXN2 and then transfected to the HEK293 cell line. Expression of recombinant bovine hyaluronidase was estimated using a hyaluronidase activity assay with gel electrophoresis. Recombinant hyaluronidase could resolve highly polymeric hyaluronic acid and also caused dispersal of the cumulus cell layer. Comparative analysis with respect to enzyme activity was carried out for the glycosylated and deglycosylated bovine sperm hyaluronidase by N-glycosidase F treatment. Finally, mutagenesis analysis revealed that among the five potential N-linked glycosylation sites, only three contributed to significant inhibition of hyaluronic activity. Recombinant bovine SPAM1 has hyaluronan degradation and cumulus oocyte complex dispersion ability, and the N-linked oligosaccharides are important for enzyme activity, providing a foundation for the commercialization of hyaluronidase.

ISSN 

1017-7825

Link 

http://dx.doi.org/10.4014/jmb.1804.04016

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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