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Lakhtin V., Lakhtin M., Aleshkin V. INTERACTION of ESTERASES

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Lakhtin V., Lakhtin M., Aleshkin V. INTERACTION of ESTERASES Lakhtin V., Lakhtin M., Aleshkin V. G.N. Gabrichevsky Research Institute for Epidemiology and Microbiology, Russia INTERACTION OF ESTERASES WITH CARBOHYDRATE-SENSITIVE REAGENTS Summary. The data on glycobiological aspects of 63 esterases (interacting with carbohydrate- sensitive proteins and other reagents) from 360 sources are systemized and presented as a table. Key words: esterases, glycosidases, lectins, glycoconjugate recognition, biotechnology, microbiology, medicine. Isolation of esterases using lectins Lectin chromatography step, as the most effective one, provided significant purification (30- 1,000 fold [f]) in the following cases of esterases: 9-O-acetylated sialate-specific esterase from the rat liver microsomes (31 f) [451]; pseudocholinesterase of fish (28 f) [424]; human milk lipase (111 f) [377], rat liver lysosomal phospholipase A1 (165-1,000 f) [440, 441]; human plasma lipoprotein lipase (85 f) [447]; bovine mammary gland microsomal alkaline phosphatase (30 f) [471]; acid phosphatase from the sunflower seeds or mammalian tissues (28-40 f) [618, 556, 601]; bovine liver cytosolic 5'-nucleotidase (33 f) [630]; soybean nucleotide-specific phosphatase (123 f) [669]; human tissue acid sphingomyelinase (54-108 f) [708, 709, 705]; human urine arylsulphatase A (119 f) [741]; mammalian liver arylsulphatase B (263-465 f) [750]; human placental N-acetylgalactosamine-6- sulphate sulphatase (64 f) [733]; nuclease SP from the spinach leaves (37 f) [810]; the differentiation-specific nuclease from the plant mesophyll cells (72 f) [808]; and fish 5'-inosinate- monophosphate hydrolase (157 f) [593]. It should be noted that in some cases the sequential chromatography (the use of coupled column sorbents including immobilized lectin and other non-lectin type sorbent or sorbents) was especially effective and suitable for isolation of the following esterases: human milk lipase, 111 f (filtration through ConA-sorbent followed by binding to cholate-sorbent, ConA-Cholate+) [377]; rat tissue lysosomal phospholipase A1 (combination of hydroxyapatite and DEAE-cellulose with ConA- sorbent (HA—DEAEC—ConA) [398], or Orange and Red dye agaroses with ConA- and phenyl- agarose [442], or hydroxyapatite with ConA-sorbent (HA—ConA), 254 f [438]); mammalian tissue arylphosphatases, 104-163 f (immobilized ConA and Blue-dye-agarose) [751, 761]; mammalian tissue iduronate- or glucuronate-2-phosphatase (immobilized ConA and Blue-dye-agarose, 119-853 f [756, 762], or immobilized ConA, Blue agarose and Green agarose [757, 1735, 1734]). A lot of examples for effective separation of esterase multiple forms using lectin sorbents can also be found in the table given below. Some esterases reveal relatively high hydrophobic properties so that ethylene glycol is needed for successful affine chromatography of enzymes using lectins [615, 622]. In some cases significant activation resulted in high purification levels of esterases using lectin sorbent alone or coupled with other type of sorbent: rat liver lysosomal phospholipase A1 (56% of activation, 165 f) [440]; rat kidney cortex lysosomal phospholipase A1 (130% of activation, 254 f) [438]; human liver N-acetylglucosmine 6-sulphate sulphatase (110% of activation, 163 f) [761]. Slight activation was observed during lectin chromatography of mammalian carboxyesterase [365], acid phosphatase [606], 5'-nucleotidase [657], or acid sphingomyelinase [709]. It should be noted that activation of esterases may depend on isoform spectrum and the concrete isozyme selected for purification using lectin chromatography [365]. Carbohydrate moiety of esterases interacting with carbohydrate-binding proteins The contents of sugars for esterases are significantly varied as it can be seen from the table: from less than 1% (w/w) for wheat germ acid phosphatase [579] up to 50-55% for lower plant phospholipase B [394] and acid phosphatases [536, 542-543, 547], or higher plant acid phosphatases [548, 620]. In majority of cases, esterases have varied N-linked glycans (see the table). Enzyme carbohydrate moiety in some cases can be associated with polysaccharides: for example, O-linked mannan of yeast acid phosphatase [539], or chitin associated with minor form of insect alkaline phosphatase [462]. Other types of carbohydrate moiety can be O-linked clusters (mucin-like) of residues of GlcNAc as in human nuclear protein tyrosine phosphatase [681], or clusters of sialic acid residues sensitive for sialic acid-binding proteins [415]. The use of such inhibitors of intracellular processing glycosylation of proteins as tunicamycin [361, 449, 443, 546, 444, 459] or swainsonine [522] allows obtaining deglycosylated esterase forms and, therefore, evaluation of the size of enzyme carbohydrate moiety. For example, rat heart monomeric extracellular phospholipase contains 12,5% sugars (7 kDa within 56 kDa) compared to intracellular form from tunicamycin-treated cells [449]; chicken or bovine adipocyte lipoprotein lipase contains 12,6 or 15,8% sugars, respectively (that means the presence of three Asn-linked glycans in the monomeric enzyme) [443, 444]. Endoglycosidase-treatments can be also used for calculation of sugar contents. Thus, rat liver microsomal carboxylesterase precursor includes 3,2% sugars sensitive for EndoH-splitting (Asn- glycans of oligomannoside type of EndoH+-enzyme form) [361, 362]; fungal lipase includes 8,1% sugars sensitive for EndoF (EndoF+-enzyme form) [369]; yeast lysophospholipase EndoH+-forms 220 and 145 kDa include 69 or 53% sugars [392]; shrimp or human liver alkaline phosphatase EndoF+-forms 50 and 75 kDa contain 22 or 33% of Asn-glycans [13, 465]; yeast acid phosphatase (EndoF+) 140 kDa includes 60% Asn-glycans of oligomannoside type [545]; leishmanial promastigote lysosomal acid phosphatase (EndoF+) 70-72 kDa includes about 30% Asn-glycans [586], and another leishmanial 3'-nucleotidase 43 kDa contains about 3% N-glycanase-sensitive glycans [671]; human protein tyrosine phosphatase 114 kDa (expressed in insect cells SF-9) includes 14% Asn-glycans of complex type [679]; murine plasmacytoma cell phosphodiesterase I (115 kDa) includes 10% N-glycanase-sensitive Asn-glycans [695]; and fungal ribonuclease (having subunits of three types) includes glycosylated polypeptides 34 and 30 kDa (21 or 8% Asn-glycans of complex type, respectively) [796]. Combinative and/or sequential glycosiadases-treatments allow deeper knowledge on glycoprotein esterases. Other endoglycosydases can be useful in study of esterases: EndoD in case of bovine 5'- nucleotidase [628], or endo-beta-galactosidase in the case of human ribonuclease [803]. In addition, the treatments of esterases with alpha-mannosidases [364, 414, 628, 780, 729], glucosidases [571, 653], beta-galactosidase [571, 628], alpha-fucosidase [571, 628], beta-N-acetylglucosaminidase [729, 628], lysozyme [729], or cellulolysin including cellulase and hemicellulase [565] were used. A lot of examples for the use of sialidase-treatments of esterases (especially for mammalian enzymes) can be found in table. As a result of such treatment, human tissue alkaline phosphatasees or rat liver 5'-nucleotidase isoforms became more cationic [497, 664] and included more restricted spectrum of forms [664]. More over, simple calculation of content of sialic acid residues in esterase treated with sialidase can be made [664]. Glycosidase-treatments can result in alteration of esterase binding to a set of lectins. For example, EndoH-treated rat brain acetylcholinesterase reveals low affinity to immobilized ligatin [422]; bovine ribonuclease-B treated with endoglycosidases loose ability to interact with ConA or WGA [787, 788]; fungal endonuclease treated with EndoH does not bind to ConA-sorbent [806]. Sialidase-treatment results in significant decreasing of the bovine acetylcholinesterase binding to immobilized WGA [414] and does not allow further precipitation of the human erythrocyte cholinesterase with this lectin [415]. However, EndoH-resistant (EndoH-) sialidase-treated form of guinea pig adipocyte lipoprotein lipase reveals increased binding ability with respect of RCA-I- 2 sorbent [446]. Interestingly, sialidase-treatment of human tissue alkaline phosphatase decreases enzyme precipitation with WGA, but significantly increases precipitation with SBA [210]. Simultaneous increasing affinity to PNA and decreasing affinity to WGA were observed for sialidase-treated frog liver acid phosphatase forms [596]. Thus contribution of sialic acid residues of O-linked glycans of mucin type into interaction between esterase molecyles and lectins takes place [210, 495, 489, 596]. Sequential glycosidase-treatments of esterases allow to study internal sugar residues in glycans, as it can be seen in the cases of bovine 5'-nucleotidase [628], or human ribonuclease and arylsulphatase [739, 803]. Additional lectin analysis of all esterase forms modified during sequentional splitting of sugars allows both control of each glycosidase type effectivity and preliminary suggestion of the whole glycan structures [628]. A set of lectins (mainly lectin sorbents) can be very effective for separation and classification (lectin typing) of esterase forms, as it was demonstrated for example for rat tissue cholinesterase [423, 426]; mammalian alkaline phosphatases [210, 479, 487, 489, 493, 497, 513, 514, 518, 522]; frog or rat liver acid phosphatases [596, 616]; human liver acid sphingomyelinase acid or neutral forms [707]; human deoxyribonuclease-I forms [767, 768]; and human urine ribonuclease-U forms [802]. The direct chemical
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