Purification of soluble Î±1,2-mannosidase from<i>Candida albicans</i>CAI-4

Mora-Montes, Héctor M.; López-Romero, Everardo; Zínker, Samuel; Ponce-Noyola, Patricia; Flores-Carreón, Arturo

doi:10.1111/j.1574-6968.2006.00093.x

Cited by 11 publications

(19 citation statements)

References 27 publications

(45 reference statements)

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“…These data suggests that the 65 kDa protein is Mns1 and also strongly indicates that the soluble and membrane-bound enzymes are immunologically related. This is in accordance with our proposal that the E-I and membrane-bound α-mannosidase are precursors to the α1,2-mannosidase E-II (Mora- Montes et al 2006Montes et al , 2008. Furthermore, unpublished data from our laboratory suggests that the membrane-bound activity is a precursor of E-I, thus confirming a close relationship between the three α-mannosidases.…”

Section: Bl21 Starsupporting

confidence: 92%

“…The modification then undergoes further processing in the ER after the transfer, with the sequential removal of the glucose units by glucosidases I and II and then removal of at least one mannose residue by the α1,2-mannosidases (Herscovics 1999a, b, Trombetta 2003. Montes et al 2004Montes et al , 2006. Previously, we have shown that the α1,2-mannosidase E-II enzyme is the product of limited proteolysis of either enzyme E-I or a membranebound α-mannosidase (Mora- Montes et al 2006Montes et al , 2008, suggesting that the three α-mannosidase isoforms are related each other.…”

mentioning

confidence: 94%

“…Montes et al 2004Montes et al , 2006. Previously, we have shown that the α1,2-mannosidase E-II enzyme is the product of limited proteolysis of either enzyme E-I or a membranebound α-mannosidase (Mora- Montes et al 2006Montes et al , 2008, suggesting that the three α-mannosidase isoforms are related each other. Indeed, this is supported by our recent report indicating that a C. albicans mns1∆ null mutant lacks both soluble and membrane-bound α-mannosidase activity (Mora-Montes et al 2007).…”

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confidence: 94%

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Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Mora-Montes

López-Romero

Zínker

et al. 2008

Mem. Inst. Oswaldo Cruz

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We have previously demonstrated that roughly 80% of the total α-mannosidase activity in Candida albicans is present as a soluble enzyme, while the remaining activity is associated with the membrane (Vázquez-Reyna et al. 1993). Further work with the soluble fraction revealed the presence of two soluble α-mannosidase isoforms, named E-I and E-II (Vázquez-Reyna et al. 1999). Both enzymes were capable of removing the α1,2-mannose residues from a M 9 N-linked mannan core, indicating that they belong to family 47 of glycosyl hydrolases (Mora- Glycosylation is the most common covalent modification of proteins found in eukaryotic cells. The glycans attached to proteins are categorised into different classes based on the type of glycosidic linkage formed between the sugar and the protein, such as N-linked, O-linked glycans and glycosylphosphatidylinositol anchors. In eukaryotic cells, N-glycosylation of proteins is initiated in the endoplasmic reticulum (ER) and completed in the Golgi complex. Formation of the N-glycosidic bond requires a lipid-linked oligosaccharide (Glc 3 Man 9 GlcNAc 2 -P-P-Dol), an asparagine (Asn) residue in an appropriate sequence within the polypeptide chain and the oligosaccharyltransferase complex that actually carries out the transfer of the oligosaccharide to the polypeptide (Kornfeld & Kornfeld 1985). The modification then undergoes further processing in the ER after the transfer, with the sequential removal of the glucose units by glucosidases I and II and then removal of at least one mannose residue by the α1,2-mannosidases (Herscovics 1999a, b, Trombetta 2003.

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Section: Bl21 Starsupporting

confidence: 92%

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confidence: 94%

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confidence: 94%

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Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Mora-Montes

López-Romero

Zínker

et al. 2008

Mem. Inst. Oswaldo Cruz

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“…The high speed supernatant was saved and used as starting material for the purification of a-mannosidase E-I, following essentially the same protocol described for the soluble enzyme from the strain CAI-4 (Mora- Montes et al 2006). Following gel chromatography on Sepharose CL6B, enzyme was recovered as a single peak of activity which was further subjected to ion exchange in DEAE.…”

Section: Resultsmentioning

confidence: 99%

“…Enzyme purification was carried out essentially as described before (Mora-Montes et al 2006). Briefly, the freeze-dried, high-speed supernatant was resuspended in 6.0 ml of buffer A and subjected to gel filtration in a column (2.7 · 57.8 cm) of Sepharose CL6B equilibrated and eluted with the same buffer.…”

Section: Organism and Culture Conditionsmentioning

confidence: 99%

Conversion of α1,2-mannosidase E-I from Candida albicans to α1,2-mannosidase E-II by limited proteolysis

Mora-Montes

López-Romero

Zínker

et al. 2007

Antonie van Leeuwenhoek

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Previous studies demonstrated the presence in Candida albicans ATCC 26555 of two soluble alpha1,2-mannosidases: E-I and E-II. In contrast, in the C. albicans CAI-4 mutant only E-I was detected and it could be processed by a membrane-bound proteolytic activity from the ATCC 26555 strain, generating an active 43 kDa polypeptide. Here, alpha1,2-mannosidase E-I from strain ATCC 26555 was purified by conventional methods of protein isolation and affinity chromatography in Concanavalin A-Sepharose 4B. Analytical electrophoresis of the purified enzyme revealed two polypeptides of 52 and 23 kDa, the former being responsible for enzyme activity as revealed by zymogram analysis. Time course proteolysis with an aspartyl protease from Aspergillus saitoi, converted alpha1,2-mannosidase E-I into an active polypeptide of 43 kDa which trimmed Man(9)GlcNAc(2), generating Man(8)GlcNAc(2) isomer B and mannose. Trimming was inhibited preferentially by 1-deoxymannojirimycin. Both, the molecular mass and the enzyme properties of the proteolytic product were identical to those described for alpha1,2-mannosidase E-II therefore supporting the notion that E-I is the precursor of E-II.

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Characterization of an extracellularly derived α-mannosidase from the liquid exudate of the sclerotia of Sclerotinia sclerotiorum (Lib.) de Bary

Liu

Wei

et al. 2015

Antonie van Leeuwenhoek

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Class I α-mannosidases play an important role in co- and post-translational N-glycosylation modification of proteins, and also in glycoprotein glycan hydrolysis. Herein, we investigated a protein named Man-41, from liquid exudate droplets secreted on the surface of developing sclerotia by Sclerotinia sclerotiorum. The protein was identified by MALDI-TOF mass spectrometry to be a α-mannosidase. The full-length open reading frame of Man-41 consists of 1581 bp, encoding 526 amino acid residues and containing a putative signal peptide at amino acid residues 1-20, and a conserved sequence at residues 50-521. Man-41 was classified into glycoside hydrolase family 47 (GH47) by clustering analysis. The catalytic residues include Glu(125), Arg(129), Asp(270), Ser(271), Glu(274), Arg(420), Glu(422), Glu(425), Glu(485), Thr(514), and Glu(515), which are conserved in all Class I α-1,2-mannosidases. Recombinant Man-41 protein had 26.67 ± 2.18 U/mg of α-mannosidase activity, about one-half of intracellular mannosidase activity of sclerotia. In conclusion, this is the first time that mannosidase has been identified in an extracellular fluid and Man-41 is also a new member of GH47 with Ca(2+)-dependent characteristics. This work lays the foundation for further study of the function of Man-41 in sclerotial development.

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Purification of soluble Î±1,2-mannosidase fromCandida albicansCAI-4

Cited by 11 publications

References 27 publications

Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Heterologous expression and biochemical characterization of an α1,2-mannosidase encoded by the Candida albicans MNS1 gene

Conversion of α1,2-mannosidase E-I from Candida albicans to α1,2-mannosidase E-II by limited proteolysis

Characterization of an extracellularly derived α-mannosidase from the liquid exudate of the sclerotia of Sclerotinia sclerotiorum (Lib.) de Bary

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