RRC ID 45029
Author Shiozaki K, Ryuzono S, Matsushita N, Ikeda A, Takeshita K, Chigwechokha PK, Komatsu M, Miyagi T.
Title Molecular cloning and biochemical characterization of medaka (Oryzias latipes) lysosomal neu4 sialidase.
Journal Fish Physiol Biochem
Abstract Glycoconjugates are known to be involved in many physiological events in vertebrates. Sialidase is one of the glycosidases, which removes sialic acid from glycoconjugates. In mammals, the properties and physiological functions of sialidases have been investigated, while there is little understanding of fish sialidase. Here, to investigate the significance of fish neu4 sialidase, neu4 gene was cloned from medaka brain mRNA and identified. Sialidase-specific motifs (GPG, YRVP and Asp-Box) were well conserved in the medaka neu4 polypeptide. Optimal pH of medaka neu4 sialidase was 4.6, but its activity was sustained even at neutral and weak alkaline pH. The neu4 considerably cleaved sialic acid from 4-methylumbelliferyl-N-acetyl-α-D-neuraminic acid and sialyllactose, but not from ganglioside and fetuin, which are good substrates for human NEU4. neu4 activity was mostly detected in mitochondria/lysosome fraction after biochemical fractionation, and indirect immunofluorescence assays revealed neu4 localization in lysosome in neu4 overexpressed cells. Next, developmental change in medaka neu4 and other sialidase mRNA levels were estimated by real-time PCR. Each sialidases showed different expression patterns in embryonic development: neu4 was up-regulated at late developmental stage in embryo, and neu3a mRNA level was quite high in 0.5 dpf. On the other hand, neu3b expression was drastically increased after hatching, suggesting that each sialidase may play a different role in embryonic development.
Volume 40(5)
Pages 1461-72
Published 2014-10-1
DOI 10.1007/s10695-014-9940-9
PMID 24744226
MeSH Amino Acid Motifs / genetics Animals Brain / metabolism* Chemical Fractionation Cloning, Molecular Computational Biology DNA Primers / genetics Fluorescent Antibody Technique, Indirect / veterinary Gene Expression Regulation, Developmental / genetics Gene Expression Regulation, Developmental / physiology* Gene Expression Regulation, Enzymologic / genetics Gene Expression Regulation, Enzymologic / physiology* HEK293 Cells Humans Hydrogen-Ion Concentration Lysosomes / enzymology* Neuraminidase / genetics* Neuraminidase / metabolism Oryzias / genetics* Oryzias / metabolism RNA, Messenger / genetics* RNA, Messenger / metabolism Real-Time Polymerase Chain Reaction / veterinary Substrate Specificity
IF 2.249
Times Cited 13
Human and Animal Cells GAKS(RCB1452)