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rdf:type | |
lifeskim:mentions | |
pubmed:dateCreated |
1995-4-28
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pubmed:abstractText |
Two acetylcholinesterases (AChE) differing in substrate and inhibitor specificities have been characterized in the medical leech (Hirudo medicinalis). A 'spontaneously-soluble' portion of AChE activity (SS-AChE) was recovered from haemolymph and from tissues dilacerated in low-salt buffer. A second portion of AChE activity was obtained after extraction of tissues in low-salt buffer alone or containing 1% Triton X-100 [detergent-soluble (DS-) AChE). Both enzymes were purified to homogeneity by affinity chromatography on edrophonium- and concanavalin A-Sepharose columns. Denaturing SDS/PAGE under reducing conditions gave one band at 30 kDa for purified SS-AChE and 66 kDa for DS-AChE. Sephadex G-200 chromatography indicated a molecular mass of 66 kDa for native SS-AChE and of 130 kDa for DS-AChE. SS-AChE showed a single peak sedimenting at 5.0 S in sucrose gradients with or without Triton X-100, suggesting that it was a hydrophylic monomer (G1). DS-AChE sedimented as a single 6.1-6.5 S peak in the presence of Triton X-100 and aggregated in the absence of detergent. A treatment with phosphatidylinositol-specific phospholipase C suppressed aggregation and gave a 7 S peak. DS-AChE was thus an amphiphilic glycolipid-anchored dimer. Substrate specificities were studied using p-nitrophenyl esters (acetate, propionate and butyrate) and corresponding thiocholine esters as substrates. SS-AChE displayed only limited variations in Km values with charged and uncharged substrates, suggesting a reduced influence of electrostatic interactions in the enzyme substrate affinity. By contrast, DS-AChE displayed higher Km values with uncharged than with charged substrates. SS-AChE was more sensitive to eserine and di-isopropyl fluorophosphate (IC50 5 x 10(-8) and 10(-8) M respectively) than DS-AChE (5 x 10(-7) and 5 x 10(-5) M.
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pubmed:commentsCorrections |
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-1125207,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-1356436,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-13726518,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-14187330,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-1901515,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-2660188,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-3024971,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-3121787,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-5432063,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-6864228,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-6875557,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-8144590,
http://linkedlifedata.com/resource/pubmed/commentcorrection/7702560-8344295
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pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical | |
pubmed:status |
MEDLINE
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pubmed:month |
Mar
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pubmed:issn |
0264-6021
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
15
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pubmed:volume |
306 ( Pt 3)
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
687-92
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pubmed:dateRevised |
2009-11-18
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pubmed:meshHeading | |
pubmed:year |
1995
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pubmed:articleTitle |
Solubilization, molecular forms, purification and substrate specificity of two acetylcholinesterases in the medicinal leech (Hirudo medicinalis).
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pubmed:affiliation |
Department of Experimental Medicine, University of Perugia, Italy.
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pubmed:publicationType |
Journal Article
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