A Conserved Acidic Motif Is Crucial for Enzymatic Activity of Protein O-Mannosyltransferases*

  title={A Conserved Acidic Motif Is Crucial for Enzymatic Activity of Protein O-Mannosyltransferases*},
  author={Mark Lommel and Andrea Schott and Thomas Jank and Verena Simone Hofmann and Sabine Strahl},
  journal={The Journal of Biological Chemistry},
  pages={39768 - 39775}
Background: Dimerization is a prerequisite for protein O-mannosyltransferase activity. Results: Transferase activity depends on an intact Asp-Glu motif located in a region crucial for acceptor binding/catalysis in both complex partners. Conclusion: Complex formation leads to the assembly of a composite catalytic center. Significance: Defining the role of complex formation is crucial for understanding the catalytic mechanism of protein O-mannosyltransferases. Protein O-mannosylation is an… 

Figures and Tables from this paper

Structure of the eukaryotic protein O-mannosyltransferase Pmt1-Pmt2 complex

Cryo-EM structures of the Saccharomyces cerevisiae Pmt1-Pmt2 complex bound to a donor and an acceptor peptide at 3.2-A resolution reveal the substrate recognition model and confirm an inverting mannosyl-transferring reaction mechanism by the enzyme complex, confirming a previously proposed evolutionary relationship between protein O-mannosylation and protein N-glycosylation.

Structure of the eukaryotic protein O-mannosyltransferase Pmt1–Pmt2 complex

Cryo-EM structures of the yeast mannosyltransferase complex Pmt1–Pmt2 bound to substrates reveal the substrate recognition model and confirm the reaction mechanism, confirming a previously proposed evolutionary relationship between protein O-mannosylation and protein N-glycosylation.

A Phenylalanine to Serine Substitution within an O-Protein Mannosyltransferase Led to Strong Resistance to PMT-Inhibitors in Pichia pastoris

This study reports the identification and characterization of a point mutation within the PpPMT2 gene that resulted in a near complete loss of PMTi sensitivity, both in terms of growth-inhibition and reduction in O-mannosylglycan site occupancy, and provides genetic evidence demonstrating that the F664 residue plays a critical role in mediating the inhibitory effects of these PMTi compounds.

Functional implications of MIR domains in protein O-mannosylation

It is shown that conserved residues in site α influence enzyme processivity of the Pmt1-Pmt2 heterodimer in vivo and a functional description of MIR domains in protein O-mannosylation is provided.

Functional Similarities between the Protein O-Mannosyltransferases Pmt4 from Bakers' Yeast and Human POMT1*

In vivo and in vitro analyses showed that general features such as protein stability of the Pmt4 variants were not significantly affected, however, the mutants proved largely enzymatically inactive, and functional and biochemical similarities between POMT1 and its orthologue from bakers' yeast PMT4 were demonstrated.

Mammalian O-mannosylation of cadherins and plexins is independent of protein O-mannosyltransferases 1 and 2

The classical and evolutionarily conserved POMT O-mannosylation pathway is essentially dedicated to α-dystroglycan and a few other proteins, whereas a novel O-mannose glycosylation process in mammalian cells is predicted to serve the large cadherin superfamily and other proteins.

Cellular Consequences of Diminished Protein O-Mannosyltransferase Activity in Baker’s Yeast

A genome-wide screen to identify Saccharomyces cerevisiae mutants with increased sensitivity towards the PMT-specific inhibitor compound R3A-5a identified the cell wall and the ER as the cell compartments affected most upon PMT inhibition, and identified Ost3, one of two homologous subunits of the oligosaccharyltransferase complexes involved in N-glycosylation, suggesting a functional link between the two pathways.

AglS, a Novel Component of the Haloferax volcanii N-Glycosylation Pathway, Is a Dolichol Phosphate-Mannose Mannosyltransferase

Relying on bioinformatics, topological analysis, gene deletion, mass spectrometry, and biochemical assays, AglS was shown to act as a dolichol phosphate-mannose mannosyltransferase, mediating the transfer of mannose from dolICHol phosphate to the tetrasaccharide corresponding to the first four subunits of the pentasaccharides N-linked to the S-layer glycoprotein.

Photoaffinity labeling of protein O-mannosyltransferases of the PMT1/PMT2 subfamily.

Protein O-mannosylation is initiated at the endoplasmic reticulum (ER) by dolichyl phosphate-mannose: protein O-mannosyltransferases (PMTs). PMTs are members of the glycosyltransferase (GT) C



Structure-Function Analysis of the Dolichyl Phosphate-Mannose: Protein O-Mannosyltransferase ScPmt1p*

It is shown that several amino acid substitutions in the conserved motifs significantly reduce ScPmt1p activity, and the invariant residues Arg-64, Glu-78, Arg-138, and Leu-408 are essential for ScPMT1p function.

Transmembrane Topology of Pmt1p, a Member of an Evolutionarily Conserved Family of Protein O-Mannosyltransferases*

This work examines the transmembrane topology of Pmt1p, an integral membrane protein of the endoplasmic reticulum, from Saccharomyces cerevisiae and proposes a seven-trans Membrane helical model for the yeast PMT1p mannosyltransferase, which is based on site-directed mutagenesis of endogenousN-glycosylation sites.

Protein-O-glycosylation in yeast: protein-specific mannosyltransferases.

The results clearly indicate that the various protein O-mannosyltransferases have different specificities for protein substrates.

PMT1, the gene for a key enzyme of protein O-glycosylation in Saccharomyces cerevisiae.

It was observed that protein O-mannosylation in the disruptant had decreased only to about 40-50%, indicating the existence of an additional transferase which had not been measured by the in vitro enzyme assay, which led to a complete loss of in vitro mannosyltransferase activity.

Protein O-glycosylation in Saccharomyces cerevisiae. Purification and characterization of the dolichyl-phosphate-D-mannose-protein O-D-mannosyltransferase.

The enzyme dolichyl-phosphate- D-mannose:protein O-D-mannosyltransferase has been solubilized from Saccharomyces cerevisiae membranes and its mannosyl transferase activity demonstrated using short peptides, indicating that the protein is a glycoprotein which may contain four carbohydrate chains.

X-ray structure of a bacterial oligosaccharyltransferase

The X-ray structure of a bacterial OST, the PglB protein of Campylobacter lari, in complex with an acceptor peptide is reported, which defines the fold of STT3 proteins and provides insight into glycosylation sequon recognition and amide nitrogen activation, both of which are prerequisites for the formation of the N-glycosidic linkage.

The PMT gene family: protein O‐glycosylation in Saccharomyces cerevisiae is vital.

The results show that protein O‐glycosylation is essential for cell wall rigidity and cell integrity and that this protein modification, therefore, is vital for Saccharomyces cerevisiae.

Protein O-mannosyltransferases participate in ER protein quality control

The results reveal previously unknown cellular roles of the Pmt1p–Pmt2p complex in connection with the ERAD machinery and show its participation in ER protein quality control.

Partial purification of a mannosyltransferase involved in the O-mannosylation of glycoproteins from Saccharomyces cerevisiae.

In general, mannosyltransferase was found to exhibit greater specificity for the L-alpha-PC derivatives in which the sn-2 position of the glycerol contained a saturated fatty acid.