Themeasurementoflimit-dextrinase(LD)(EC3.2.1.142)ingrainsamplessuchasbarley,wheatorricecanbeproblematicforanumberofreasons.TheintrinsicLDactivityinthesesamplesisextremelylowandtheyoftencontainalimit-dextrinaseinhibitorand/orhighlevelsofreducingsugars.LDalsoexhibitstransglycosylationactivitythatcancomplicatethemeasurementofitshydrolyticactivity.AminormodificationtotheindustrialstandardLimit-Dextrizymetablettestissuggestedheretoovercomethistransglycosylationissue.Inaddition,twonewsubstratesaredescribedthatcanbeadoptedforuseinanauto-analyserformat.4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-6³-α-D-maltotriosyl-maltotrioside(BzCNPG₃G₃,Hexachrom)isnotsusceptIBLetotransglycosylationandservesamiablyasaroutinequantitativeassaytoolwiththepotentialtorunkineticassaysduetothelowpK_a(∼5.5)ofthechromogenicmoietywhile4,6-O-benzylidene-4-methylumbelliferyl-β-6³-α-D-maltotriosyl-maltotrioside(BzMUG₃G₃,Hexafluor)wasfoundtobesusceptibletotransglycosylationwithLD.ItisanticipatedthatHexafluormayfindextensiveuseinapplicationswherehighsensitivityisrequiredsuchashighthroughputscreeningstudies.

Specificandhighlysensitivecolourimetricandfluorometricsubstratemixtureshavebeenpreparedforthemeasurementofpullulanaseandlimit-dextrinaseactivityandassaysemployingthesesubstrateshavebeendeveloped.ThesemixturescompriseThermostableα-andβ-glucosidasesandeither4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-maltotriosyl(1-6)α-maltotrioside(BzCNPG₃G₃,1)asacolourimetricsubstrateor4,6-O-benzylidene-4-methylumbelliferyl-β-maltotriosyl(1-6)α-maltotrioside(BzMUG₃G₃,2)asafluorometricsubstrate.Hydrolysisofsubstrates1and2byexo-actingenzymessuchasamyloglucosidase,β-amylaseandα-glucosidaseispreventedbythepresenceofthe4,6-O-benzylidenegrouponthenon-reducingendD-glucosylresidue.Thesubstratesarenothydrolysedbyanyα-amylasesstudied,(includingthosefromAspergillusnigerandporcinepancreas)andareresistanttohydrolysisbyPseudomonassp.isoamylase.Onhydrolysisbypullulanase,the2-chloro-4-nitrophenyl-β-maltotrioside(3)or4-methylumbelliferyl-β-maltotrioside(4)liberatedisimmediatelyhydrolysedtoD-glucoseand2-chloro-4-nitrophenolor4-methylumbelliferone.ThereactionisterminatedbytheadditionofaweakalkalinesolutionleADIngtotheformationofphenolateionsinsolutionwhoseconcentrationcanbedeterminedusingeitherspectrophotometricorfluorometricanalysis.Theassayprocedureissimpletouse,specific,accurate,robustandreadilyadaptedtoautomation.

β-1,3-D-Glucansarebiologicalresponsemodifierswithpotenteffectsontheimmunesystem.Anumberofreceptorsarethoughttoplayaroleinmediatingtheseresponses,includingmurineDectin-1,whichwerecentlyidentifiedasaβ-glucanreceptor.Inthisstudywedescribethecharacterizationofthehumanhomologueofthisreceptorandshowthatitisstructurallyandfunctionallysimilartothemousereceptor.Thehumanβ-glucanreceptorisatypeIItransmembranereceptorwithasingleextracellularcarbohydraterecognitiondomainandanimmunoreceptortyrosineactivationmotifinitscytoplasmictail.Thehumanβ-glucanreceptoriswidelyexpressedandfunctionsasapatternrecognitionreceptor,recognizingavarietyofβ-1,3-and/orβ-1,6-linkedglucansaswellasintactyeast.Incontrasttothemurinereceptor,thehumanreceptormRNAisalternativelyspliced,resultingintwomajor(AandB)andsixminorisoforms.Thetwomajorisoformsdifferbythepresenceofastalkregionseparatingthecarbohydraterecognitiondomainfromthetransmembraneregionandaretheonlyisoformsthatarefunctionalforβ-glucanbinding.ThehumanreceptoralsobindsT-lymphocytesatasitedistinctfromtheβ-glucanbindingsite,indicatingthatthisreceptorcanrecognizebothendogenousandexogenousligands.