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PredicateObject
rdf:type
lifeskim:mentions
pubmed:dateCreated
1995-4-28
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.
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
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
0264-6021
pubmed:author
pubmed:issnType
Print
pubmed:day
15
pubmed:volume
306 ( Pt 3)
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
687-92
pubmed:dateRevised
2009-11-18
pubmed:meshHeading
pubmed:year
1995
pubmed:articleTitle
Solubilization, molecular forms, purification and substrate specificity of two acetylcholinesterases in the medicinal leech (Hirudo medicinalis).
pubmed:affiliation
Department of Experimental Medicine, University of Perugia, Italy.
pubmed:publicationType
Journal Article