Title :
C-Mannosylation of human
RNase 2 is an intracellular process performed by a variety of cultured cells
Abstract :
- C2-alpha-Mannosyltryptophan was discovered in RNase 2 from human urine , representing a novel way of attaching carbohydrate to a protein
- Here, we have addressed two questions related to the biosynthesis of this modification: (i) is C-mannosylation part of the normal intracellular biosynthetic route, and (ii) how general is it, i.e. which organisms perform this kind of glycosylation
- To answer the first question, RNase 2 , which is identical to the eosinophil-derived neurotoxin , was isolated from intracellular stores of cultured human HL-60 cells
- The enzyme was C-mannosylated at Trp-7 , showing that the modification occurs intracellularly, before secretion of the protein
- The second question was investigated by immunological and chemical analysis of RNase 2 purified from the supernatant of transiently transformed cells from different organisms
- This revealed that C-mannosylation occurs in cells from man, green monkey, pig, mouse, and hamster
- The observation that pig kidney cells contain the machinery for C-mannosylation of Trp-7 of human RNase 2 but that the homologous RNase from porcine kidney is not a substrate, since it does not contain a tryptophan at position 7 , strongly suggests that C-mannosylated proteins other than RNase 2 exist
- Recombinant RNase 2 isolated from insect cells, plant protoplasts, and Escherichia coli was not C-mannosylated
- These results not only form the basis for further studies on the biochemical aspects of C-mannosylation but also have implications for the choice of cells for production of recombinant glycoproteins