Biogenesis of the glycosome in Trypanosoma brucei: the synthesis, translocation and turnover of glycosomal polypeptides.

Abstract

Glycosomes, the microbodies of Trypanosoma brucei, contain a number of enzymes involved in glucose and glycerol metabolism. The biogenesis of three of these enzymes has been studied. Aldolase, D-glyceraldehyde-3-phosphate dehydrogenase and NAD-linked glycerol-3-phosphate dehydrogenase are all synthesized in the cytosol on free rather than on membrane-bound polysomes. In vitro, as well as in vivo, these polypeptides are synthesized at their mature size, and no evidence was found for any processing upon entry into the glycosomes. Continuous and pulse-chase labelling experiments with procyclic trypomastigotes revealed that the enzymes have a half-life in the cytosol of -3 min or less, and then turn over rapidly in the glycosomes, with half-lives as short as 30 min.

Biogenesis of glycosomes of Trypanosoma brucei: an in vitro model of 3-phosphoglycerate kinase import.

“…2A). No larger precursors were observed, consistent with the previous observation that these polypeptides are synthesized at their mature sizes (6).…”

“…Thus, glycosome assembly would require insertion of proteins across the organelle membrane posttranslationally. In the cases examined thus far, in vitro translation of mRNA yields GPs with the same molecular weights as the mature products inside the glycosome (6), suggesting that the import may not involve proteolytic processing. Thus, this process appears similar to the biogenesis of other microbodies, such as the peroxisomes of yeast and mammals (7)(8)(9) and the glyoxysomes of plants (10), and differs from the biogenesis of mitochondria and chloroplasts where posttranslational protein import generally involves proteolytic cleavage of specific leader sequences (11)(12)(13).…”

“…brucei glycosomes contain 10% of the total cellular protein (4) but no nucleic acids (5). It has been assumed that the genes encoding glycosomal proteins (GPs) are located in the nucleus and the products of these genes are synthesized in the cytoplasm on free polysomes (6). Thus, glycosome assembly would require insertion of proteins across the organelle membrane posttranslationally.…”

The cytosolic and glycosomal isoenzymes of glyceraldehyde‐3‐phosphate dehydrogenase in Trypanosoma brucei have a distant evolutionary relationship

“…Since glycosomal proteins do not undergo any detectable form of proteolytic processing or secondary modification upon entry into the organelle [52], the topogenic signal must be present in the structure of the mature protein. We proposed originally that the high positive charge would play an essential role in the import, since it was common to all glycosomal enzymes analyzed, whereas the C-terminal extension was not [6, 10, 53 -571.…”

In vivo import of firefly luciferase into the glycosomes of Trypanosoma brucei and mutational analysis of the C-terminal targeting signal.

“…An evolutionary relationship between glycosomes and peroxisomes was proposed based on their structural and functional resemblance (Borst, 1986;Michels and Opperdoes, 1991) and the fact that, like peroxisomal proteins in higher eukaryotes, glycosomal proteins are encoded by nuclear genes, synthesized on free polyribosomes, and imported into the glycosomes without proteolytic modifications (Hart et al, 1987). Most peroxisomal proteins contain at their C-terminals the tripeptide serine-lysine-leucine (SKL) or a closely analogous tripeptide, which is necessary for the import of these proteins into peroxisomes (Gould et al, 1989).…”