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Kelley W Moremen - One of the best experts on this subject based on the ideXlab platform.

  • Family 47 α‐Mannosidases in N‐Glycan Processing
    Methods in Enzymology, 2020
    Co-Authors: Steven W Mast, Kelley W Moremen
    Abstract:

    Abstract α‐Mannosidases in eukaryotic cells are involved in both glycan biosynthetic reactions and glycan catabolism. Two broad families of enzymes have been identified that cleave terminal mannose linkages from Asn‐linked oligosaccharides (Moremen, 2000), including the Class 1 Mannosidases (CAZy GH family 47 (Henrissat and Bairoch, 1996)) of the early secretory pathway involved in the processing of N‐glycans and quality control and the Class 2 Mannosidases (CAZy family GH38 [Henrissat and Bairoch, 1996]) involved in glycoprotein biosynthesis or catabolism. Within the Class 1 family of α‐Mannosidases, three subfamilies of enzymes have been identified (Moremen, 2000). The endoplasmic reticulum (ER) α1,2‐mannosidase I (ERManI) subfamily acts to cleave a single residue from Asn‐linked glycans in the ER. The Golgi α‐mannosidase I (GolgiManI) subfamily has at least three members in mammalian systems (Herscovics et al., 1994; Lal et al., 1994; Tremblay and Herscovics, 2000) involved in glycan maturation in the Golgi complex to form the Man5GlcNAc2 processing intermediate. The third subfamily of GH47 proteins comprises the ER degradation, enhancing α‐mannosidase‐like proteins (EDEM proteins) (Helenius and Aebi, 2004; Hirao et al., 2006; Mast et al., 2005). These proteins have been proposed to accelerate the degradation of misfolded proteins in the lumen of the ER by a lectin function that leads to retrotranslocation to the cytosol and proteasomal degradation. Recent studies have also indicated that ERManI acts as a timer for initiation of glycoprotein degradation via the ubiquitin‐proteasome pathway (Hosokawa et al., 2003; Wu et al., 2003). This article discusses methods for analysis of the GH47 α‐Mannosidases, including expression, purification, activity assays, generation of point mutants, and binding studies by surface plasmon resonance.

  • Molecular cloning and expression of an α-mannosidase gene in Mycobacterium tuberculosis
    Microbial Pathogenesis, 2020
    Co-Authors: Carlos A. Rivera-marrero, Jeffrey D. Ritzenthaler, Jesse Roman, Kelley W Moremen
    Abstract:

    Abstract Mannose is a major component of glycolipids and glycoproteins of the cell envelope of M. tuberculosis ( Mtb ). However, the enzymes involved in the biosynthesis and catabolism of mannoIsylated glycans are largely unknown. We demonstrate α-mannosidase activity towards the fluorescent substrate 4-methylumberlliferyl-α-D-mannopyranoside (4MU-Man) in cell lysates of attenuated and virulent Mtb bacilli, with two-fold higher activity in the virulent strain Erdman. Mannosidase activity was optimal at pH 6.5, was not inhibited by deoxymannojirimycin (dMNJ), was mildly inhibited by swainsonine (SW) and stimulated two-fold by EDTA. GenBank BLAST analysis for sequences homologous to eukaryotic α-Mannosidases revealed a 3.6 kb putative gene (Rv0648) in Mtb cosmid SCY20H10 (Acc# z92772), with strong homology (48%) to the rat ER/cytosolic α-mannosidase and containing signature sequences of class 2 Mannosidases. By RT-PCR, gene Rv0648 was found differentially expressed, with lower expression during growth in A549 pneumocyte cultures. Gene Rv0648 was cloned, expressed in E. coli , and α-mannosidase activity in cell lysates determined. Expression of αMan-pET in E. coli cells resulted in an eight-fold increase in mannosidase activity toward 4-MU-Man, upon IPTG induction. Partial purification of the histidine-tagged Mtb mannosidase by metal chelation affinity chromatography, and analysis by SDS-PAGE, showed a protein with the predicted m.w. of 137.5 kDa. Enzyme assays of the column fractions showed α-mannosidase activity toward synthetic aryl-mannose substrates, in fractions enriched in the recombinant Mtb mannosidase. These results demonstrate that gene Rv0648 encodes an active α-mannosidase in Mtb .

  • substrate recognition and catalysis by gh47 α Mannosidases involved in asn linked glycan maturation in the mammalian secretory pathway
    Proceedings of the National Academy of Sciences of the United States of America, 2016
    Co-Authors: Yong Xiang, Khanita Karaveg, Kelley W Moremen
    Abstract:

    Maturation of Asn-linked oligosaccharides in the eukaryotic secretory pathway requires the trimming of nascent glycan chains to remove all glucose and several mannose residues before extension into complex-type structures on the cell surface and secreted glycoproteins. Multiple glycoside hydrolase family 47 (GH47) α-Mannosidases, including endoplasmic reticulum (ER) α-mannosidase I (ERManI) and Golgi α-mannosidase IA (GMIA), are responsible for cleavage of terminal α1,2-linked mannose residues to produce uniquely trimmed oligomannose isomers that are necessary for ER glycoprotein quality control and glycan maturation. ERManI and GMIA have similar catalytic domain structures, but each enzyme cleaves distinct residues from tribranched oligomannose glycan substrates. The structural basis for branch-specific cleavage by ERManI and GMIA was explored by replacing an essential enzyme-bound Ca2+ ion with a lanthanum (La3+) ion. This ion swap led to enzyme inactivation while retaining high-affinity substrate interactions. Cocrystallization of La3+-bound enzymes with Man9GlcNAc2 substrate analogs revealed enzyme–substrate complexes with distinct modes of glycan branch insertion into the respective enzyme active-site clefts. Both enzymes had glycan interactions that extended across the entire glycan structure, but each enzyme engaged a different glycan branch and used different sets of glycan interactions. Additional mutagenesis and time-course studies of glycan cleavage probed the structural basis of enzyme specificity. The results provide insights into the enzyme catalytic mechanisms and reveal structural snapshots of the sequential glycan cleavage events. The data also indicate that full steric access to glycan substrates determines the efficiency of mannose-trimming reactions that control the conversion to complex-type structures in mammalian cells.

  • family 47 α Mannosidases in n glycan processing
    Methods in Enzymology, 2006
    Co-Authors: Steven W Mast, Kelley W Moremen
    Abstract:

    Abstract α‐Mannosidases in eukaryotic cells are involved in both glycan biosynthetic reactions and glycan catabolism. Two broad families of enzymes have been identified that cleave terminal mannose linkages from Asn‐linked oligosaccharides (Moremen, 2000), including the Class 1 Mannosidases (CAZy GH family 47 (Henrissat and Bairoch, 1996)) of the early secretory pathway involved in the processing of N‐glycans and quality control and the Class 2 Mannosidases (CAZy family GH38 [Henrissat and Bairoch, 1996]) involved in glycoprotein biosynthesis or catabolism. Within the Class 1 family of α‐Mannosidases, three subfamilies of enzymes have been identified (Moremen, 2000). The endoplasmic reticulum (ER) α1,2‐mannosidase I (ERManI) subfamily acts to cleave a single residue from Asn‐linked glycans in the ER. The Golgi α‐mannosidase I (GolgiManI) subfamily has at least three members in mammalian systems (Herscovics et al., 1994; Lal et al., 1994; Tremblay and Herscovics, 2000) involved in glycan maturation in the Golgi complex to form the Man5GlcNAc2 processing intermediate. The third subfamily of GH47 proteins comprises the ER degradation, enhancing α‐mannosidase‐like proteins (EDEM proteins) (Helenius and Aebi, 2004; Hirao et al., 2006; Mast et al., 2005). These proteins have been proposed to accelerate the degradation of misfolded proteins in the lumen of the ER by a lectin function that leads to retrotranslocation to the cytosol and proteasomal degradation. Recent studies have also indicated that ERManI acts as a timer for initiation of glycoprotein degradation via the ubiquitin‐proteasome pathway (Hosokawa et al., 2003; Wu et al., 2003). This article discusses methods for analysis of the GH47 α‐Mannosidases, including expression, purification, activity assays, generation of point mutants, and binding studies by surface plasmon resonance.

  • energetics of substrate binding and catalysis by class 1 glycosylhydrolase family 47 α Mannosidases involved inn glycan processing and endoplasmic reticulum quality control
    Journal of Biological Chemistry, 2005
    Co-Authors: Khanita Karaveg, Kelley W Moremen
    Abstract:

    Abstract Nascent glycoproteins are subject to quality control in the lumen of the endoplasmic reticulum (ER) where they can either be effectively folded with the aid of a collection of ER chaperones or they can be targeted for disposal in a process known as ER-associated degradation. Initiation of the ER disposal process involves selective trimming of N-glycans by ER α-mannosidase I and subsequent recognition by the ER degradation-enhancing α-mannosidase-like protein family of lectins, both members of glycosylhydrolase family 47. The kinetics and energetics of substrate binding and catalysis by members of this family were investigated here by the analysis of wild type and mutant forms of human ER α-mannosidase I. The contributions of several amino acid residues and an enzyme-associated Ca2+ ion to substrate binding and catalysis were demonstrated by a combination of surface plasmon resonance and enzyme kinetic analyses. One mutant, E330Q, shown previously to alter general acid function within the catalytic site, resulted in an enzyme that possessed increased glycan binding affinity but compromised glycan hydrolysis. This mutant protein was used in a series of glycan binding studies with a library of mannose-containing ligands to examine the energetics of Man9GlcNAc2 substrate interactions. These studies provide a framework for understanding the nature of the unusual substrate interactions within the family 47 Mannosidases involved in glycan maturation and ER-associated glycoprotein degradation.

Annette Herscovics - One of the best experts on this subject based on the ideXlab platform.

  • stimulation of erad of misfolded null hong kong α1 antitrypsin by golgi α1 2 Mannosidases
    Biochemical and Biophysical Research Communications, 2007
    Co-Authors: Nobuko Hosokawa, Linda O Tremblay, Kazuhiro Nagata, Annette Herscovics
    Abstract:

    Abstract Terminally misfolded or unassembled proteins are degraded by the cytoplasmic ubiquitin-proteasome pathway in a process known as ERAD (endoplasmic reticulum-associated protein degradation). Overexpression of ER α1,2-mannosidase I and EDEMs target misfolded glycoproteins for ERAD, most likely due to trimming of N -glycans. Here we demonstrate that overexpression of Golgi α1,2-mannosidase IA, IB, and IC also accelerates ERAD of terminally misfolded human α1-antitrypsin variant null (Hong Kong) (NHK), and mannose trimming from the N -glycans on NHK in 293 cells. Although transfected NHK is primarily localized in the ER, some NHK also co-localizes with Golgi markers, suggesting that mannose trimming by Golgi α1,2-Mannosidases can also contribute to NHK degradation.

  • structure and function of class i α1 2 Mannosidases involved in glycoprotein synthesis and endoplasmic reticulum quality control
    Biochimie, 2001
    Co-Authors: Annette Herscovics
    Abstract:

    Abstract Class I α1,2-Mannosidases (glycosylhydrolase family 47) are conserved through eukaryotic evolution. This protein family comprises three subgroups distinguished by their enzymatic properties. The first subgroup includes yeast ( Saccharomyces cerevisiae ) and human α1,2-Mannosidases of the endoplasmic reticulum that primarily form Man 8 GlcNAc 2 isomer B from Man 9 GlcNAc 2 . The second subgroup includes mammalian Golgi α1,2-Mannosidases, as well as enzymes from insect cells and from filamentous fungi, that trim Man 9 GlcNAc 2 to Man 8 GlcNAc 2 isomers A and/or C intermediates toward the formation of Man 5 GlcNAc 2 . Yeast and mammalian proteins of the third subgroup have no enzyme activity with Man 9 GlcNAc 2 as substrate. The members of subgroups 1 and 3 participate in endoplasmic reticulum quality control and promote proteasomal degradation of misfolded glycoproteins. The yeast endoplasmic reticulum α1,2-mannosidase has served as a model for structure-function studies of this family. Its structure was determined by X-ray crystallography as an enzyme-product complex. It consists of a novel (αα) 7 barrel containing the active site that includes essential acidic residues and calcium. The structures of the subgroup 1 human endoplasmic reticulum α1,2-mannosidase and of a subgroup 2 fungal α1,2-mannosidase were determined by molecular replacement. Comparison of the enzyme structures is providing some insight into the reasons for their different specificities.

  • α Mannosidases involved in n glycan processing show cell specificity and distinct subcompartmentalization within the golgi apparatus of cells in the testis and epididymis
    European Journal of Cell Biology, 1999
    Co-Authors: Suleiman A Igdoura, Annette Herscovics, Kelley W Moremen, Carlos R Morales, Louis Hermo
    Abstract:

    The Golgi apparatus is enriched in specific enzymes involved in the maturation of carbohydrates of glycoproteins. Among them, α-Mannosidases IA, IB and II are type II transmembrane Golgi-resident enzymes that remove mannose residues at different stages of N-glycan maturation. α-Mannosidases IA and IB trim Man9GlcNAc2 to Man5GlcNAc2, while α-mannosidase II acts after GlcNAc transferase I to remove two mannose residues from GlcNAcMan5GlcNAc2 to form GlcNAcMan3GlcNAc2 prior to extension into complex N-glycans by Golgi glycosyltransferases. The objective of this study is to examine the expression as well as the subcellular localization of these Golgi enzymes in the various cells of the male rat reproductive system. Our results show distinct celland region-specific expression of the three Mannosidases examined. In the testis, only α-mannosidase IA and II were detectable in the Golgi apparatus of Sertoli and Leydig cells, and while α-mannosidase IB was present in the Golgi apparatus of all germ cells, only the Golgi apparatus of steps 1-7 spermatids was reactive for α-mannosidase IA. In the epididymis, principal cells were unreactive for α-mannosidase II, but they expressed α-mannosidase IB in the initial segment and caput regions, and α-mannosidase IA in the corpus and cauda regions. Clear cells expressed n-mannosidase II in all epididymal regions, and α-mannosidase IB only in the caput and corpus regions. Ultrastructurally, α-mannosidase IB was localized mainly over cis saccules, α-mannosidase IA was distributed mainly over trans saccules, and α-mannosidase II was localized mainly over medial saccules of the Golgi stack. Thus, the cell-specific expression and distinct Golgi subcompartmental localization suggest that these three α-Mannosidases play different roles during N-glycan maturation.

  • substrate specificities of recombinant murine golgi α1 2 Mannosidases ia and ib and comparison with endoplasmic reticulum and golgi processing α1 2 Mannosidases
    Glycobiology, 1998
    Co-Authors: Peng Pang, Annette Herscovics, Sandeep Kalelkar, Pedro A Romero, Kelley W Moremen
    Abstract:

    : The catalytic domains of murine Golgi alpha1,2-Mannosidases IA and IB that are involved in N-glycan processing were expressed as secreted proteins in P.pastoris . Recombinant Mannosidases IA and IB both required divalent cations for activity, were inhibited by deoxymannojirimycin and kifunensine, and exhibited similar catalytic constants using Manalpha1,2Manalpha-O-CH3as substrate. Mannosidase IA was purified as a 50 kDa catalytically active soluble fragment and shown to be an inverting glycosidase. Recombinant Mannosidases IA and IB were used to cleave Man9GlcNAc and the isomers produced were identified by high performance liquid chromatography and proton-nuclear magnetic resonance spectroscopy. Man9GlcNAc was rapidly cleaved by both enzymes to Man6GlcNAc, followed by a much slower conversion to Man5GlcNAc. The same isomers of Man7GlcNAc and Man6GlcNAc were produced by both enzymes but different isomers of Man8GlcNAc were formed. When Man8GlcNAc (Man8B isomer) was used as substrate, rapid conversion to Man5GlcNAc was observed, and the same oligosaccharide isomer intermediates were formed by both enzymes. These results combined with proton-nuclear magnetic resonance spectroscopy data demonstrate that it is the terminal alpha1, 2-mannose residue missing in the Man8B isomer that is cleaved from Man9GlcNAc at a much slower rate. When rat liver endoplasmic reticulum membrane extracts were incubated with Man9GlcNAc2, Man8GlcNAc2was the major product and Man8B was the major isomer. In contrast, rat liver Golgi membranes rapidly cleaved Man9GlcNAc2to Man6GlcNAc2and more slowly to Man5GlcNAc2. In this case all three isomers of Man8GlcNAc2were formed as intermediates, but a distinctive isomer, Man8A, was predominant. Antiserum to recombinant mannosidase IA immunoprecipitated an enzyme from Golgi extracts with the same specificity as recombinant mannosidase IA. These immunodepleted membranes were enriched in a Man9GlcNAc2to Man8GlcNAc2-cleaving activity forming predominantly the Man8B isomer. These results suggest that Mannosidases IA and IB in Golgi membranes prefer the Man8B isomer generated by a complementary mannosidase that removes a single mannose from Man9GlcNAc2.

  • The Saccharomyces cerevisiae Processing α1,2-Mannosidase Is an Inverting Glycosidase
    Biochemical and Biophysical Research Communications, 1995
    Co-Authors: F. Lipari, B.j. Goursalin, Annette Herscovics
    Abstract:

    Abstract The α1,2-mannosidase from Saccharomyces cerevisiae , which removes one specific α1,2-linked mannose residue from Man 9 GlcNAc 2 , is a member of the Class 1 α1,2-mannosidase family conserved from yeast to mammals. Although Class 1 α1,2-Mannosidases are essential for the maturation of N-linked oligosaccharides in mammalian cells, nothing is known about their mechanism of action. The availability of sufficient quantities of recombinant yeast α1,2-mannosidase and its homology with the mammalian enzymes make it a good model to study the catalytic mechanism of this family of α1,2-Mannosidases. The stereochemical course of hydrolysis of Man 9 GlcNAc by the yeast enzyme was followed by proton nuclear magnetic resonance spectroscopy. It was observed that β-D-mannose is released from the oligosaccharide substrate, thereby demonstrating that the enzyme is of the inverting type.

Michael A Curtis - One of the best experts on this subject based on the ideXlab platform.

  • characterization of the and Mannosidases of porphyromonas gingivalis
    Journal of Bacteriology, 2013
    Co-Authors: Minnie Rangarajan, Joseph Aduseopoku, Ahmed Hashim, Nikolay Paramonov, Michael A Curtis
    Abstract:

    Mannose is an important sugar in the biology of the Gram-negative bacterium Porphyromonas gingivalis. It is a major component of the oligosaccharides attached to the Arg-gingipain cysteine proteases, the repeating units of an acidic lipopolysaccharide (A-LPS), and the core regions of both types of LPS produced by the organism (O-LPS and A-LPS) and a reported extracellular polysaccharide (EPS) isolated from spent culture medium. The organism occurs at inflamed sites in periodontal tissues, where it is exposed to host glycoproteins rich in mannose, which may be substrates for the acquisition of mannose by P. gingivalis. Five potential Mannosidases were identified in the P. gingivalis W83 genome that may play a role in mannose acquisition. Four Mannosidases were characterized in this study: PG0032 was a -mannosidase, whereas PG0902 and PG1712 were capable of hydrolyzingp-nitrophenyl -D-mannopyranoside. PG1711 and PG1712 were -1i3 and-1i2 Mannosidases, respectively. No enzyme function could be assigned to PG0973. -1i6 mannobiose was not hydrolyzed by P. gingivalis W50. EPS present in the culture supernatant was shown to be identical to yeast mannan and a component of the medium used for culturing P. gingivalis and was resistant to hydrolysis by Mannosidases. Synthesis of O-LPS and A-LPS and glycosylation of the gingipains appeared to be unaffected in all mutants. Thus, - and -Mannosidases of P. gingivalis are not involved in the harnessing of mannan/mannose from the growth medium for these biosynthetic processes. P. gingivalis grown in chemically defined medium devoid of carbohydrate showed reduced -mannosidase activity (25%), suggesting these enzymes are environmentally regulated.

  • characterization of the α and β Mannosidases of porphyromonas gingivalis
    Journal of Bacteriology, 2013
    Co-Authors: Minnie Rangarajan, Joseph Aduseopoku, Ahmed Hashim, Nikolay Paramonov, Michael A Curtis
    Abstract:

    Mannose is an important sugar in the biology of the Gram-negative bacterium Porphyromonas gingivalis. It is a major component of the oligosaccharides attached to the Arg-gingipain cysteine proteases, the repeating units of an acidic lipopolysaccharide (A-LPS), and the core regions of both types of LPS produced by the organism (O-LPS and A-LPS) and a reported extracellular polysaccharide (EPS) isolated from spent culture medium. The organism occurs at inflamed sites in periodontal tissues, where it is exposed to host glycoproteins rich in mannose, which may be substrates for the acquisition of mannose by P. gingivalis. Five potential Mannosidases were identified in the P. gingivalis W83 genome that may play a role in mannose acquisition. Four Mannosidases were characterized in this study: PG0032 was a β-mannosidase, whereas PG0902 and PG1712 were capable of hydrolyzing p-nitrophenyl α-d-mannopyranoside. PG1711 and PG1712 were α-1→3 and α-1→2 Mannosidases, respectively. No enzyme function could be assigned to PG0973. α-1→6 mannobiose was not hydrolyzed by P. gingivalis W50. EPS present in the culture supernatant was shown to be identical to yeast mannan and a component of the medium used for culturing P. gingivalis and was resistant to hydrolysis by Mannosidases. Synthesis of O-LPS and A-LPS and glycosylation of the gingipains appeared to be unaffected in all mutants. Thus, α- and β-Mannosidases of P. gingivalis are not involved in the harnessing of mannan/mannose from the growth medium for these biosynthetic processes. P. gingivalis grown in chemically defined medium devoid of carbohydrate showed reduced α-mannosidase activity (25%), suggesting these enzymes are environmentally regulated.

David R Rose - One of the best experts on this subject based on the ideXlab platform.

  • human lysosomal α Mannosidases exhibit different inhibition and metal binding properties
    Protein Science, 2009
    Co-Authors: Meenakshi Venkatesan, Douglas A Kuntz, David R Rose
    Abstract:

    Two structurally-related members of the lysosomal mannosidase family, the broad substrate specificity enzyme human lysosomal α-mannosidase (hLM, MAN2B1) and the human core α-1, 6-specific mannosidase (hEpman, MAN2B2) act in a complementary fashion on different glycosidic linkages, to effect glycan degradation in the lysosome. We have successfully expressed these enzymes in Drosophila S2 cells and functionally characterized them. hLM and hEpman were significantly inhibited by the class II α-mannosidase inhibitors, swainsonine and mannostatin A. We show that three pyrrolidine-based compounds designed for selective inhibition of Golgi α-mannosidase II (GMII) exhibited varying degrees of inhibition for hLM and hEpman. While these compounds inhibited hLM and GMII similarly, they inhibited hEpman to a lesser extent. Further, the two lysosomal α-Mannosidases also show differential metal dependency properties. This has led us to propose a secondary metal binding site in hEpman. These results set the stage for the development of selective inhibitors to members of the GH38 family, and, henceforth, the further investigation of their physiological roles.

  • functionalized pyrrolidine inhibitors of human type ii α Mannosidases as anti cancer agents optimizing the fit to the active site
    Bioorganic & Medicinal Chemistry, 2008
    Co-Authors: Helene Fiaux, Douglas A Kuntz, David R Rose, Daniela Hoffman, Robert C Janzer, Sandrine Gerberlemaire, Lucienne Juilleratjeanneret
    Abstract:

    Refining the chemical structure of functionalized pyrrolidine-based inhibitors of Golgi α- mannosidase II (GMII) to optimize binding affinity provided a lead molecule that demonstrated nanomolar competitive inhibition of α-Mannosidases II and an optimal fit in the active site of Drosophila GMII by X-ray crystallography. Esters of this lead compound also inhibited the growth of human glioblastoma and brain-derived endothelial cells more than the growth of non-tumoral human fibroblasts, suggesting their potential for anti-cancer therapy.

  • comparison of kifunensine and 1 deoxymannojirimycin binding to class i and ii alpha Mannosidases demonstrates different saccharide distortions in inverting and retaining catalytic mechanisms
    Biochemistry, 2003
    Co-Authors: Niket Shah, Douglas A Kuntz, David R Rose
    Abstract:

    Mannosidases are key enzymes in the eukaryotic N-glycosylation pathway. These enzymes fall into two broad classes (I and II) and are characteristically different in catalytic mechanism, sequence, and structure. Kifunensine is an alkaloid that is a strong inhibitor against class I α-Mannosidases but is only a weak inhibitor against class II α-Mannosidases. In this paper, the 1.80 A resolution crystal structure of kifunensine bound to Drosophila melanogaster Golgi α-mannosidase II (dGMII) is presented. Kifunensine adopts a 1,4B boat conformation in the class II dGMII, which contrasts the 1C4 chair conformation seen in class I human endoplasmic reticulum α1,2 mannosidase (hERMI, PDB 1FO2). The observed conformations are higher in conformational energy than the global minimum 4C1 conformation, although the conformation in hERMI is closer to the minimum, as supported by an energy calculation. Differing conformations of 1-deoxymannojirimycin were also observed:  a 4C1 and 1C4 conformation in dGMII and hERMI, re...

Igor Polikarpov - One of the best experts on this subject based on the ideXlab platform.

  • insights into the structure and function of fungal β Mannosidases from glycoside hydrolase family 2 based on multiple crystal structures of the trichoderma harzianum enzyme
    FEBS Journal, 2014
    Co-Authors: Alessandro S. Nascimento, Alexander M. Golubev, Joao R.enato C. Muniz, Ricardo Aparicio, Igor Polikarpov
    Abstract:

    Hemicellulose is an important part of the plant cell wall biomass, and is relevant to cellulosic ethanol technologies. β-Mannosidases are enzymes capable of cleaving nonreducing residues of β-d-mannose from β-d-mannosides and hemicellulose mannose-containing polysaccharides, such as mannans and galactomannans. β-Mannosidases are distributed between glycoside hydrolase (GH) families 1, 2, and 5, and only a handful of the enzymes have been structurally characterized to date. The only published X-ray structure of a GH family 2 mannosidase is that of the bacterial Bacteroides thetaiotaomicron enzyme. No structures of eukaryotic Mannosidases of this family are currently available. To fill this gap, we set out to solve the structure of Trichoderma harzianum GH family 2 β-mannosidase and to refine it to 1.9-A resolution. Structural comparisons of the T. harzianum GH2 β-mannosidase highlight similarities in its structural architecture with other members of GH family 2, reveal the molecular mechanism of β-mannoside binding and recognition, and shed light on its putative galactomannan-binding site. Database Coordinates and observed structure factor amplitudes have been deposited with the Protein Data Bank (4CVU and 4UOJ). The T. harzianum β-mannosidase 2A nucleotide sequence has GenBank accession number BankIt1712036 GeneMark.hmm KJ624918.

  • Insights into the structure and function of fungal β-Mannosidases from glycoside hydrolase family 2 based on multiple crystal structures of the Trichoderma harzianum enzyme
    The FEBS journal, 2014
    Co-Authors: Alessandro S. Nascimento, Alexander M. Golubev, Joao R.enato C. Muniz, Ricardo Aparicio, Igor Polikarpov
    Abstract:

    UNLABELLED: Hemicellulose is an important part of the plant cell wall biomass, and is relevant to cellulosic ethanol technologies. β-Mannosidases are enzymes capable of cleaving nonreducing residues of β-d-mannose from β-d-mannosides and hemicellulose mannose-containing polysaccharides, such as mannans and galactomannans. β-Mannosidases are distributed between glycoside hydrolase (GH) families 1, 2, and 5, and only a handful of the enzymes have been structurally characterized to date. The only published X-ray structure of a GH family 2 mannosidase is that of the bacterial Bacteroides thetaiotaomicron enzyme. No structures of eukaryotic Mannosidases of this family are currently available. To fill this gap, we set out to solve the structure of Trichoderma harzianum GH family 2 β-mannosidase and to refine it to 1.9-Å resolution. Structural comparisons of the T. harzianum GH2 β-mannosidase highlight similarities in its structural architecture with other members of GH family 2, reveal the molecular mechanism of β-mannoside binding and recognition, and shed light on its putative galactomannan-binding site. DATABASE: Coordinates and observed structure factor amplitudes have been deposited with the Protein Data Bank (4CVU and 4UOJ). The T. harzianum β-mannosidase 2A nucleotide sequence has GenBank accession number BankIt1712036 GeneMark.hmm KJ624918.