Purification and characterization of a ribonuclease from Aspergillus giganteus IFO 5818, the gigantin. Immunological and enzymic comparison with alphasarcin.
A protein has been isolated from a culture medium of Aspergillus giganteus IFO 5818 (gigantin) and purified by ion-exchange chromatography successively on DEAE-cellulose and carboxymethyl-cellulose, and gel-filtration chromatography on Biogel P10. With a high purity, gigantin was found to be a non-glycosylated basic protein with a relative molecular mass of 17000 +/- 200 determined in PAGE/SDS. Gigantin was able to digest the synthetic homopolymers of nucleic acids poly(A), poly(I), poly(C) and poly(U). The catalytic action has an optimal pH around 7.0, an optimal temperature at 45-55 degrees C and can be inhibited by cations. Gigantin activity, analyzed as its capacity to hydrolyze RNA from yeast, was comparable to that of alphasarcin, a similar biochemical protein produced by the strain A. giganteus MDH 18894. A study of the substrate specificity for alphasarcin indicated a preference for poly(A) and poly(I), while gigantin had greater activity on poly(C) and poly(U). The cross reaction of gigantin with a rabbit antiserum to alphasarcin suggests a high sequence similarity between both proteins. However, gigantin is immunologically distinguishable from alphasarcin as alphasarcin antiserum detects epitopes in alphasarcin that are not present in gigantin.
Salvarelli
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S
Departmento de Investigación, Hospital Ramón y Cajal, Madrid, Spain.
Muñoz
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Conde
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FP
D001230:Aspergillus / Q000201:enzymology* / Q000254:growth & development; D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D004722:Endoribonucleases; D005656:Fungal Proteins / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007118:Immunoassay; D005779:Immunodiffusion; D007700:Kinetics; D008970:Molecular Weight; D011131:Polyribonucleotides / Q000378:metabolism; D012260:Ribonucleases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D013379:Substrate Specificity
D005656:Fungal Proteins; D011131:Polyribonucleotides; C014801:alpha-sarcin; D004722:Endoribonucleases; D012260:Ribonucleases; C090042:gigantin