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SIALYLTRANSFERASES IN RAT BRAIN: REACTION KINETICS, PRODUCT ANALYSES, AND MULTIPLICITIES OF ENZYME SPECIES
Author(s) -
Ng S.S.,
Dain J. A.
Publication year - 1977
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1977.tb06511.x
Subject(s) - glycolipid , fetuin , sialyltransferase , endogeny , lactosylceramide , biochemistry , enzyme , glycoprotein , gel electrophoresis , biosynthesis , substrate (aquarium) , chemistry , biology , ecology
— The incorporation of NeuNAc from CMP‐NeuNAc into endogenous glycolipids and glyco‐proteins, and exogenously added GM 1a (monosialoganglioside) and desialylated fetuin (DS‐fetuin) was studied with particulate preparations from 11 to 15 day old rat cerebra. The apparent +K++ m values of the enzyme systems for the different substrates, assayed with 0.5 mg enzyme protein, were: CMP‐NeuNAc, 0.13 m m (same with endogenous and exogenous glycolipid and glycoprotein substrates); GM 1a , 0.20 m m ; DS‐fetuin, 0.15 m m (or 1.2 m m in terms of acceptor sites). The activities, expressed as nmoles NeuNAc incorporated per 0.5 mg enzyme protein per 30 min incubation at 37°C and pH 6.3, were 0.094, 0.039, 0.17 and 0.64 with the endogenous glycolipids, endogenous glycoproteins, exogenous GM 1a and exogenous DS‐fetuin, respectively. Incorporation into endogenous glycolipids was mainly in GM 3 , while exogenously added GM 1a was converted to GD 1a . Incorporation into endogenous glycoproteins yields about 20 sialoglycopolypeptides on SDS‐polyacrylamide gel electrophoresis. Neura‐minidase pretreatment of the particulate enzyme preparation decreased sialylation of the higher molecule weight polypeptides but increased sialylation of the lower molecule weight species. The sialyltransferase activity with the endogenous glycolipid substrates was more heat resistant than the activities with exogenous GM 1a . Since more than 60% of the endogenous glycolipid activity was due to the conversion of lactosylceramide to GM 3 , the sialyltransferase responsible for this reaction appears to be different from the one that acts on GM 1a . This was supported by the observation that exogenously added GM 1a did not diminish the incorporation of NeuNAc into endogenous lactosylceramide. These two glycolipid sialyltransferase activities were distinguishable from the glycoprotein sialyltransferase activity since exogenous DS‐fetuin did not compete with either the endogenous or the exogenous glycolipids for CMP‐NeuNAc.