I.U.B.:3.1.27.5
Pancreaticribonuclease(RNaseI)catalyzescleavageofthephosphodiesterbondbetweenthe5"-riboseofanucleotideandthephosphategroupattachedtothe3"-riboseofanadjacentpyrimidinenucleotideforminga2",3"-cyclicphosphatewhichmaythenbehydrolyzedtothecorresponding3"-nucleosidephosphate.RibonucleaseAhasamolecularweightof13,700daltons.ItoperatesinanoptimumpHrangeof7.0-7.5.
RibonucleaseBhasamolecularweightof14,700±300daltons.ItisaglycoproteinwhichpossessesanaminoacidcompositionindistinguishablefromthatofRNaseA.Itcontains6residuesofmannoseand2residuesofN-acetylglucosaminepermolecule.ItisaglycosylatedderivativeofRNaseA.
RibonucleaseisinhibitedbyheavymetalionsanditiscompetitivelyinhibitedbyDNA.SinceMolecularBiologyGradeRNaseA(ProductCode:RPDF)isessentiallyfreeofDNaseandproteaseactivities,thisproductisusefulinremovingRNAfromDNAinnucleicacidworkandwhereotherenzymesareusedorwhereintactproteinsmustberecovered.
StABIlity/Storage:MolecularBiologyGradeproduct(ProductCode:RPDF)isstableatleast2yearsat2-8°Cor-20°C.OthergradesofRNaseAandRNaseBarestable2-3yearsat2-8°C.R,RB,andRAF:Storeat2-8°Candprotectfrommoisture.ProductCode:RASE:Storeat-20°Ctomaintainmonomericform.ProductCode:RPDF:Storeat2-8°Corat-20°C.
UnitDefinition:OneKalnitskyUnitcausesanincreaseinabsorbanceof1.0at260nmat37°CandpH5.0whenyeastribosomalRNAishydrolyzedtoacidsolubleoliognucleotides.OneKunitzunitequals50Worthingtonunits.
TechnicalNotes:Specialcareshouldbegiventohandlingoftheenzymebecauseofitsaffinityforglasssurfaces.Theenzymeremainsactivebutaggregatesuponlyophilizationandinsolutionattemperatures≥2°Catlowionicstrength.HeatingsolutionsofRNaseAtoinactivateDNasemaynotbesatisfactorysinceRNaseactivitymaybelostifprecipitateformationoccursandheattreatedDNasemayreactivateovertime.ProductCode:RPDFissuitableassuppliedforapplicationsrequiringminimalDNaseandproteaselevelsandneedsnofurthertreatment.ProductCode:RAFcanbeusedwithouttreatmentinsomeapplications.Toheat-treatRAF,use10mMacetatepH5.0withorwithout15mMCaCl2for15minutesat100°Corlongerat80°C.ProductmayprecipitateifheatedatneutralpH.HeattreatmentofProductCode:RASEwillprecipitateproductduetothepresenceofphosphate.
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