Granulosain I, a Cysteine Protease Isolated from Ripe Fruits of Solanum granuloso -leprosum (Solanaceae)

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 24,746 Da (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 0.6 mM) 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.

Kinetic studies on recombinant stem bromelain

“…Hence, there are only minor but noticeable differences in the specificity of recBM and cBM to the substrates. The data obtained in this study is comparable to the kinetic data reported for cysteine proteases including stem bromelain [20,24] granulosain [21], ascepalin [25], hieronymain II [26] and penduliforain I [27]. Based on kinetic data obtained in this study, it can be inferred that the kinetic behavior of recBM (containing His 6x tag, signal and propeptide) is not much different to that of mature cBM.…”

“…In pH range of 5 -8, the enzymes have hydrogen bond between the thiol and imidazole functional group (Cys-His) that is critical for catalytic activity [21]. The order of susceptibility for the hydrolysis of synthetic substrates and the substrate specificity showed by recBM and cBM are apparent.…”

Kinetics studies with fruit bromelain (Ananas comosus) in the presence of cysteine and divalent ions

“…At pH 5-8, the enzyme has hydrogen bond between thiol and imidazole functional group (i.e. Cys-His) and is critical for catalytic activity (Valle et al 2008). Furthermore, the observed fruit bromelainpH profile was consistent with the thiol proteases of papain family, which display a wide profile of maximum activity around neutral pH (Rowan et al 1990).…”

“…Kinetic studies with stem bromelain by Valle et al (2008) suggested that at pH 5-8, the enzyme has hydrogen bond between thiol and imidazole functional group (i.e. Cys-His) and is critical for its catalytic activity.…”

Transcriptomic Analysis of Acclimation to Temperature and Light Stress in Saccharina latissima (Phaeophyceae)

“…Interestingly the induction of two genes annotated as different cysteine proteases, cysteine proteinase 2 and papain family cysteine protease containing protein was observed in S. latissima under excessive light and high temperature. Cysteine proteases are involved in intracellular protein degradation, they respond to different internal and external stimuli and are able to provide up to 90% of the proteolytic activity [82], [83]. Moreover, several studies showed that cysteine protease activity is a key event in programmed cell death or apoptosis in plants and different phytoplankton species [84]–[87].…”