Mariela Bruno

Publications (2)2.08 Total impact

• Article: Purification and characterization of hieronymain III. Comparison with other proteases previously isolated from Bromelia hieronymi Mez.

  Mariela A Bruno, Sebastián A Trejo, Néstor O Caffini, Laura M I López
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  ABSTRACT: 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.
  The Protein Journal 12/2008; 27(7-8):426-33. · 1.04 Impact Factor
• Article: Granulosain I, a Cysteine Protease Isolated from Ripe Fruits of Solanum granuloso-leprosum (Solanaceae)

  Diego Vallés, Mariela Bruno, Laura M. I. López, Néstor O. Caffini, Ana María B. Cantera
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  ABSTRACT: A new cysteine peptidase (Granulosain I) was isolated from ripe fruits of Solanum granuloso-leprosum Dunal (Solanaceae) by means of precipitation with organic solvent and cation exchange chromatography. The enzyme showed a single band by SDS-PAGE, its molecular mass was 24,746Da (MALDI-TOF/MS) and its isoelectric point was higher than 9.3. It showed maximum activity (more than 90%) in the pH range 7–8.6. Granulosain I was completely inhibited by E-64 and activated by the addition of cysteine or 2-mercaptoethanol, confirming its cysteinic nature. The kinetic studies carried out with PFLNA as substrate, showed an affinity (Km 0.6mM) slightly lower than those of other known plant cysteine proteases (papain and bromelain). The N-terminal sequence of granulosain I (DRLPASVDWRGKGVLVLVKNQGQC) exhibited a close homology with other cysteine proteases belonging to the C1A family.
  The Protein Journal 04/2008; 27(5):267-275. · 1.04 Impact Factor