β-1,3-Glucanase and chitinase activities in soybean root nodules

Abstract

Summary β-1,3-Glucanase and chitinase specific activities were measured in soybean nodules and the adjoining root tissue during nodule ontogeny. Activities of both enzymes exhibited a transient pattern in root nodules with a maximum rate occurring following the decline in nitrogen fixation rate at 28 days post inoculation. β-1,3-Glucanase and chitinase levels were higher in the adjacent root tissue than in the nodules. β-1,3-Glucanase activity was optimal at a pH of 5.3 and an incubation temperature of 37 °C. The product formation increased linearly with time. TLC analysis of the hydrolytic products from the nodule reaction mixture revealed the presence of at least seven oligomers. ‘Western’ immunoblot analysis showed the nodule glucanases cross-reacting with antisera raised against virus-inducible tobacco ( Nicotiana tabacum L.) protein PR-1 (17 kDa), tobacco basic protein PR-2 (21 kDa), lima bean ( Phaseolus vulgaris L.) acidic protein PL1 (18 kDa), pinto bean basic protein PR-4d (21 kDa) and tobacco (36 kDa) and cowpea ( Vigna unguiculata L.) (36 kDa) PR-proteins. Chitinase was heat-labile and sensitive to mercury but was protease-resistant. A non-denaturing polyacrylamide gel activity stain revealed the presence of at least four prominent chitinolytic isoforms in nodule extract as well as in root tissue. ‘Western’ immunoblot analysis showed cross-reactivitiy of nodule chitinases with an anti-vacuolar bean leaf chitinase.