Multiple forms of β-Hexosaminidase in ripening tomato (Lycopersicum Esculantum L.)

The specific activity of β-hexosaminidase in tomato fruits increased during ripening with a peak at climacteric stage. The enzyme was purified from climacteric fruit for studying its properties. Two major isoforms (I and II) of β-hexosaminidase were separated by ion-exchange chromatography on DEAE-Sephadex A-50, with a percent abundance of 57 and 37, respectively, a third one being very minor. The isoforms I and II of β-hexosaminidase were individually subjected to gel permeation chromatography on Sephadex G-200 followed by PAGE. The pH optimum was 5.0 and 4.6 for isoforms -I and -II, respectively. The temperature optimum was 47°C for isoform I, which had a thermostability of 72% at 57°C for 15 min. Isoform II displayed a temperature optimum of 47–57°C, with a thermostability of 100% under similar conditions. Both the isoforms of β-hexosaminidase were inhibited by Hg²⁺, Zn²⁺ and Cu²⁺. The Km values for p-nitrophenyl-b-N-acetyl glucosaminopyranoside were 1.6 and 1.1mM, and molecular weights were > 94 & 64 kDa, respectively, for the isoforms-I and -II.