Cloning and Expression of cDNA for a Human Gal(β1‐3)GalNAc α2,3‐Sialyltransferase from the CEM T‐Cell Line
1997; Wiley; Volume: 247; Issue: 2 Linguagem: Inglês
10.1111/j.1432-1033.1997.00558.x
ISSN1432-1033
AutoresValérie Giordanengo, Sylvie Bannwarth, C. Laffont, Vincent Van Miecem, Anne Harduin‐Lepers, Philippe Delannoy, Jean‐Claude Lefebvre,
Tópico(s)Peptidase Inhibition and Analysis
ResumoComplementary DNA encoding a human Gal(β1‐3)GalNAc α2,3‐sialyltransferase type II (hST3Gal II) was cloned from a CEM T‐cell cDNA library using a 23‐base oligonucleotide probe. The sequence of this probe was established on the basis of a slightly divergent sialylmotif L that was obtained by polymerase chain reaction with degenerate oligonucleotide primers based on the conserved sialylmotif L of mammalian Gal(β1‐3)GalNAc α2,3‐sialyltransferases. It was thus confirmed that a short oligonucleotide probe may be sensitive and highly specific. The nucleotide and amino acid sequences of hST3Gal II show, respectively, 56.3% and 49.3% similarity to hST3Gal I [Kitagawa, H. & Paulson, J. C. (1994) J. Biol. Chem. 269 , 17872–17878] and 88.1% and 93.7% similarity to inurine ST3Gal II [Lee, Y. C., Kojima, N., Wada, E., Kurosawa, N., Nakaoka, T., Hamamoto, T. & Tsuji, S. (1994) J. Biol. Chem. 269 , 10028–10033]. hST3Gal II mRNA was highly expressed in heart, liver, skeletal muscle and various lymphoid tissues but not in brain and kidney. A soluble form of hST3Gal II expressed in COS‐7 cells was tested in vitro for substrate specificity and kinetic properties. Asialofetuin and asialo‐bovine submaxillary mucin appeared better substrates for hST3Gal II than for its murine counterpart as previously reported [Kojima, N., Lee, Y.‐C., Hamamoto, X, Kurosawa, N. & Tsuji, S. (1994) Biochemistry 33 , 5772–57761. In previous studies, we have shown hyposialylation of O ‐glycans attached to two major lymphocyte CD43 and CD45 cell surface molecules in human‐immunodeficiency‐virus‐1(H1V‐1) –infected T‐cell lines. Since comparable levels of hST3Gal 1 and hST3Gal II mRNA and enzymatic activity were observed in parental and HIV‐1‐infected CEM T‐cell lysates, the sialylation defect associated with HIV infection of this cell line is probably due to a mechanism different from a simple altered catalytic activity of these sialyltransferases.
Referência(s)