A Novel Method for Simultaneous Production of Two Ribosome-Inactivating Proteins, α-MMC and MAP30, from Momordica charantia L

Abstract

Alpha-momorcharin (α-MMC) and momordica anti-HIV protein (MAP30) from Momordica charantia L. have been confirmed to possess anti-tumor and anti-virus activities. Traditional purification methods of these two ribosome-inactivating proteins (RIPs) were separate which was time consuming and cost effective as well as low efficient. In order to obtain sufficient samples for researches, a strategy combining ion-exchange and gel filtration chromatography was developed and optimized in this study. Using this novel purification method, averagely 1162 mg of α-MMC and 535 mg of MAP30 were obtained from 400 g of Momordica charantia L seeds. The homogeneities of them were assessed by electrophoresis analysis. Determination of molecular weights of α-MMC and MAP30 were 28.585 kDa and 29.094 kDa by MALDI-TOF/TOF and pI were 9.02 and 9.12, respectively. The single glycoproteins were identified by Periodate-Schiff's base (PAS) and the saccharide content was tested to be 1.25% and 1.1% by anthrone-sulfuric acid method. Biological activities were evidenced by their ability to inhibit proliferation of lung adenocarcinoma A549 cell and to convert supercoiled plasmid pUC18 into relaxed forms. Finally, we also found that both two RIPs exhibited no superoxide dismutase (SOD) activity.

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“…We inferred that one way MAP30 may take part in WSSV and V. alginolyticus defense is by regulating humoral immune reactions. However, the activity of SOD in the serum of P. clarkii significantly declined following injection of MAP30, but MAP30 exhibited no SOD activity in vitro as well as in previous reports [46]. We propose that MAP30 may have an antioxidant capacity, and as expected, the protein was determined by the scavenging activity of hydroxyl radicals.…”

Cytotoxicity mechanism of α-MMC in normal liver cells through LRP1 mediated endocytosis and JNK activation

“…The a-MMC protein sample was kindly given by Professor Yanfa Meng, from the College of Life Sciences of Sichuan University, and it was prepared as described by Li et al (2009) andMeng et al (2014). Briefly, the preparation methods are as follows: supernatant extraction from bitter melon seeds was carried out with acetate buffer (50 mM, pH 5.0), ammonium sulphate fractionation (35-70%), MacroCap SP chromatography with a 0-0.2 M NaCl linear gradient in 20 mM phosphate buffer (PB, pH 7.0) and Superdex 75 chromatography with 20 mM PB containing 0.15 M NaCl, pH 7.0.…”

The Plant Ribosome-Inactivating Proteins Play Important Roles in Defense against Pathogens and Insect Pest Attacks

“…For example, trichosanthin (TCS), luffin α, luffin β, Mirabilis antiviral protein (MAP), camphorin, barley ( Hordeum vulgare ) translation inhibitor and saporin (from soapwort, S. officinalis ) are type I RIPs. Meng et al (2014) combined ion-exchange and gel filtration chromatography methods to isolate two RIPs, α-MMC and an anti-HIV protein (MAP30) from bitter melon. The α-MMC and MAP30 are also type I RIPs with molecular weights of 28.585 and 29.094 kDa, respectively.…”