Intact α-1,2-endomannosidase is a typical type II membrane protein

Abstract

Rat endomannosidase is a glycosidic enzyme that catalyzes the cleavage of di-, tri-, or tetrasaccharides (Glc(1-3)Man), from N-glycosylation intermediates with terminal glucose residues. To date it is the only characterized member of this class of endomannosidic enzymes. Although this protein has been demonstrated to localize to the Golgi lumenal membrane, the mechanism by which this occurs has not yet been determined. Using the rat endomannosidase sequence, we identified three homologs, one each in the human, mouse, and rat genomes. Alignment of the four encoded protein sequences demonstrated that the newly identified sequences are highly conserved but differed significantly at the N-terminus from the previously reported protein. In this study we have cloned two novel endomannosidase sequences from rat and human cDNA libraries, but were unable to amplify the open reading frame of the previously reported rat sequence. Analysis of the rat genome confirmed that the 59- and 39-termini of the previously reported sequence were in fact located on different chromosomes. This, in combination with our inability to amplify the previously reported sequence, indicated that the N-terminus of the rat endomannosidase sequence previously published was likely in error (a cloning artifact), and that the sequences reported in the current study encode the intact proteins. Furthermore, unlike the previous sequence, the three ORFs identified in this study encode proteins containing a single N-terminal transmembrane domain. Here we demonstrate that this region is responsible for Golgi localization and in doing so confirm that endomannosidase is a type II membrane protein, like the majority of other secretory pathway glycosylation enzymes.

A Pooling Genome-Wide Association Study Combining a Pathway Analysis for Typical Sporadic Parkinson’s Disease in the Han Population of Chinese Mainland

“…Scheme of the gene FLJ43860 region under investigation and relative pairwise LD plot for the sPD sample. High r 2 values are represented in red, whereas smaller r 2 values are represented by a gradual decrease of color intensity glycans, which is a type II membrane protein, like the majority of other secretory pathway glycosylation enzymes [23].…”

Heterologous Expression, Purification, and Characterization of an α-Mannosidase Belonging to Glycoside Hydrolase Family 99 ofShewanella amazonensis

“…To date, GH99 endo--mannosidases from rat 1,[3][4][5]16,17) and human 18,19) have been characterized. In early processing of eukaryotic N-linked oligosaccharides of glycoproteins, glucosidases I and II and -mannosidase I sequentially trim the Glc 3 Man 9 GlcNAc 2 structure.…”

Triple arginines in the cytoplasmic tail of endomannosidase are not essential for type II membrane topology and Golgi localization

“…The mechanism by which the enzyme is retained in the Golgi apparatus is currently unknown. Cloning of endomannosidase [24][25][26] revealed a high interspecies amino acid sequence identity [26]. Endomannosidases share particular features, i.e., three consecutive positively charged amino groups close to the N terminus followed by a single stretch of hydrophobic amino acids (residues 10-25) and conserved amino acid regions close to the C terminus [26].…”