Mannosidase purification

S. Singh, M. Scigelova, and D. H. G. Crout, Glycosidase-catalysed synthesis of mannobioses by the reverse hydrolysis activity of alpha-mannosidase Partial purification of alpha-mannosidases from almond meal, limpets and Aspergillus niger, Tetrahedron Asymmetry, 11 (2000) 223-229. 

Since then it has been established that emulsin is a mixture of enzymes comprising in addition to /3-glucosidase28 the enzymes a-galactosidase, a-mannosidase, /3-glucuronidase and /3-(N-acetyl)-glucosaminidase. Concerning the process of purification of the Rohferment and the mode of differentiation of the constituent enzymes of emulsin, the most informa-... 

J. Schweden, G. Legler, and E. Bause, Purification and characterization of a neutral processing mannosidase from calf liver acting on (Man)9(GlcNAc)2 oligosaccharides, Eur. J. Biochem., 157 (1986) 563-570. 

Purification of a-D-Mannosidase from Jack-bean Meal39... 

As discussed under Purification and pH and Stability [see Sections 11,4 (p. 409) and 11,6 (p. 413)], a-D-mannosidase is unstable at low pH values unless Zn2+ is added. In the following experiments employing a partially purified, jack-bean meal preparation (see Table V, stage 3 p. 410) to which Zn2+ had not been added, the enzyme was pre-incubated at 37° and pH 5 before assay at the same pH with p-nitrophenyl a-D-mannopyranoside as the sub-... 

As the Zn2+ required for catalytic activity could be retained during purification, a-D-mannosidase may be classified as a metalloenzyme. The enzyme seems to be completely specific for Zn2+ as the activating cation. 

An amidrazone (58) derived from 5-amino-5-deoxy-L-fuconolactam was found to inhibit a recombinant human a-L-fucosidase with a Krvalue of 820 nmol/1 [ 111 ]. A simple synthesis of 1,5-dideoxy-1,5-imino-D-arabinitol (59), previously prepared by Ganem et al. [49] as a potential mannosidase inhibitor, was applied to the affinity purification of a-L-fucosidase from bovine kidney by an improved method and the characterization of the enzyme thus obtained [112]. The relatively low affinity of this compound to the enzyme (Kj 2.2 pmol/1 at pH 7) compared to 1-deoxyfuconojirimycin (51) turned out to be advantageous in terms of enzyme recovery and yield. Structurally related, suitably protected 5-amino-5-deoxy-D-arabinopyranose (60), was coupled with a N-acetyl-6-deoxy-6-thio-D-glucosaminide (61) to give a stable thioglycoside (62) [113]. 

Ichishima, E., Arai, M., Shigematsu, Y., Kumagai, H., and Sumida-Tanaka, R. (1981). Purification of an acidic a-D-mannosidase from Aspergillus saitoi and specific cleavage of 1,2-a-D-mannosidic linkage in yeast mannan. Biochim. Biophys. Acta, 658,45-53. 

Jelinek-Kelly, S., and Herscovics, A. (1988). Glycoprotein biosynthesis in Saccharomyces cervisiae. Purification of the a-mannosidase which removes one specific mannose residue from Man9GlcNAc. J. Biol. Chem., 263, 14757-14763. 

Oku, H., Hase, S., and Ikenaka, T. (1991). Purification and characterization of neutral a-mannosidase that is activated by Co2+ from Japanese quail oviduct. J. Biochem. (Tokyo), 110,29-34. 

Lipari, F., and Herscovics, A. (1994). Production, purification and characterization of recombinant yeast processing al,2-mannosidase. Glycobiology, 4, 697-702. 

There have been only a few kinetic studies on plant /3-D-manno-sidases, because of the apparently limited distribution of this enzyme in various species, and also because of the paucity of attempts at purification. The enzyme from malted barley has been purified 41-fold by fractionation with ammonium sulfate, and chromatography302 on Biogel P-100, DEAE-cellulose, and CM-cellulose. Some properties of /3-D-mannosidases from various tissues are summarized in Table II. 

Helenius A, Aebi M. Roles of A-linked glycans in the endoplasmic reticulum. Ann. Rev. Biochem. 2004 73 1019-1049. Tulsiani DRP, Touster O. The purification and characterization of mannosidase la from rat liver Golgi membranes. J. Biol. Chem. 1988 263(11) 5408-5417. 

Castanospermine, isolated from the seeds of Castanospermum australe and from dried pods of Alexa leiopetala [6], is another polyhydroxylated indolizidine v th a potent, competitive and reversible inhibitory effect on several glycosidases. For this reason it has been largely used as a glycosidase identifier and as an auxiliary in the purification of glycosidases and mannosidases [120-125]. Castanospermine was also used in two other... 

The isolation, purification, and characterization of oc-D-mannosidase from cultures of Aspergillus flavus has been described.The enzyme (pH optimum 4.2-4.5) was isolated from the growth medium of the mold at the beginning of the stationary phase of growth and partly purified by ion-exchange chromatography. The effect of temperature and cations on the activity of the enzyme and the effect of pH on its stability were examined for 4-nitrophenyl a-D-mannopyranoside 1.4 mM, the value of the sedimentation constant is. y o = 4.5 S, and the partial specific volume q = 0.725 cm g" ). 

A glycoprotein which has been isolated from the cell walls of baker s yeast was shown to be a D-mannan-protein complex. After immobilization by covalent attachment to macroporous agarose, this was used in the purification of an a-D-mannosidase from jack-bean meal. 

D-Mannanase (j3-D-mannosidases) purified from the germinated seeds of Trifolium repens by a procedure that included chromatography on hydroxyapatite, gel filtration on acrylamide/agarose (Ultragel 5/4) and preparative polyacrylamide gel electrophoresis. The final purification has been completely resolved into two ]3-D-mannanases (I and II) with distinct specificities (see p. 468). 

Two forms of a-D-mannosidase from yellow wax beans Phaseolus vulgaris) have been purified by immunoadsorption. A high degree of immunological identity has been demonstrated between a-D-mannosidase I and II from P. vulgaris. This made possible the purification of both forms of the enzyme by chromatography on an immunoadsorbent prepared from anti-fa-D-mannosidase I) antiserum. 

Purification of detergent-solubilized la antigens Isolation of a-o-mannosidase from jack bean by affinity chromatography Preparation of specific anti-(unc 54-myosin) antibodies by immunoadsorption Affinity chromatography of glycosidases... 

A single passage of a crude extract of human placenta through a column of concanavalin A immobilized on macroporous agarose effected the purification of an a-D-mannosidase by 300-fold. The nature of the interaction with concanavalin A indicated that the a-D-mannosidase is a glycoprotein. 

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