A 96-KDa Glycyrrhizin-Binding Protein (gp96) from Soybeans Acts as a Substrate for Case in Kinase II, and Is Highly Realted to Lipoxygenase3

• Published in: Journal of biochemistry (Tokyo) 1995-12, Vol.118 (6), p.1145-1150
• Main Authors: Ohtsuki, Kenzo, Nakamura, Sanae, Shimoyama, Yoshihito, Shibata, Daisuke, Munakata, Hiroshi, Yoshiki, Yumiko, Okubo, Kazuyoshi
• Format: Article
• Language: English
• Subjects: casein kinase II; glycyrrhizin-binding protein; lipoxygenase 3; soybean; triterpenoid saponin
• ISSN: 0021-924X
• DOI: 10.1093/oxfordjournals.jbchem.a125000

A 96-kDa glycyrrhizin (GL)-binding protein (gp96) was purified to apparent homogeneity from an aqueous extract of soybeans by means of successive DEAE-cellulose column chromatography, gel filtration on Superdex 200pg, GL-affinity column chromatography, and ion-exchange chromatography on a Mono S column (HPLC). The protein was identified as a GL-binding protein since it specifically binds to [3H] GA. Moreover, it is a lipoxygenase (an enzyme that catalyzes the oxygenation of unsaturated fatty acids) since (i) it displays lipoxygenase (LOX) activity at pH 6.5; (ii) it is recognized on Western blot analysis by antibodies against LOX-1 and LOX-2; and (iii) the sequence of the N-terminal 21 amino acid residues (SNDVYLPRDEAFGHLKSSDFL) of a 42-kDa fragment (p42) proteolytically generated from gp96 is identical to a sequence of soybean LOX-3. In addition, GL, glycyrrhetinic acid (GA) and soyasaponin βg slightly inhibited LOX activity of the purified gp96 fraction, whereas oGA (a GA derivative) greatly inhibited its activity. Furthermore, CK-II catalyzed phosphorylation of gp96 was stimulated significantly by GL at doses between 1 and 10 μM, but this phosphorylation was inhibited completely by 50 μM GL. All these results taken together suggest that (i) gp96 purified from soybeans as a GL-binding protein belongs to the LOX family; and (ii) triterpenoid saponins, including GL, are involved in the regulation of the activities of CK-II and LOXs in plants, such as soybeans and roots of liquorice, which contain large quantities of saponins.