Purification and characterization of hieronymain III. Comparison with other proteases previously isolated from Bromelia hieronymi Mez.
A new proteolytic enzyme, named hieronymain III, has been purified by ion-exchange chromatography from unripe fruits of Bromelia hieronymi Mez. The new peptidase belongs to the cysteine catalytic type, as well as hieronymain I and II, the other two peptidases previously isolated from this species. Hieronymain III showed optimum alkaline pH range (8.6-9.3) and the molecular mass (MALDI-TOF) was 23713 Da. The N-terminal sequence (AVPQSIDWRRYGAVTTSRNQG) exhibited a higher percentage identity with hieronymain II (93%) than with hieronymain I (71%). The three peptidases showed notable differences on synthetic substrates degradation: whereas hieronymain III was the only one able to hidrolyze Z-Arg-Arg-p-nitroanilide, hieronymain I and II could degrade Z-Phe-Arg-p-nitroanilide; on the other hand, PFLNA was only split by hieronymain I. Finally, the three proteases showed different preferences on N-alpha-CBZ-p-nitrophenyl aminoacid ester substrates. From a biotechnological point of view, cleavage specificity differences are significant enough to use these enzymes as potential tools in that area.
Bruno
Mariela A
MA
Laboratorio de Investigación de Proteínas Vegetales (LIPROVE), Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, C.C. 711, La Plata B1900AVW, Argentina. brunomariela@biol.unlp.edu.ar
Trejo
Sebastián A
SA
Caffini
Néstor O
NO
López
Laura M I
LM
D000581:Amidohydrolases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D000595:Amino Acid Sequence; D031231:Bromelia / Q000201:enzymology*; D002852:Chromatography, Ion Exchange; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004950:Esterases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D005638:Fruit / Q000201:enzymology; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D010940:Plant Proteins; D004950:Esterases; D003546:Cysteine Endopeptidases; D000581:Amidohydrolases; C019665:amidase