Partial Purification and Characterization of Proteases in Tobacco Leaf and Callus

Ammonium sulfate fra.ctionation, gel permeation and cation-exchange column chromatography were employed for panial purification of proteases from leaf laminae and callus tissues of Samsun NN tobacco (Nicotiana tabacum L). The predominant proteases in the leaf and callus are acidic sulfhydryl proteases which are activated by 2-mercaptoethanol and ethylenediamine tetraacetic acid, completely inhibited- by iodoacetic acid, and partially inhibited by phenylmethane sulfonyl fluoride and pepstatin A. With hemoglobin and tobaccl:l Fraction I protein as substrates, leaf and callus proteases showed a pH optimum of 5. However, specific activity was significandy higher in the callus than in the leaf. Tobacco proteases digested hemoglobin more effectively than Fraction I protein and showed the least activity with casein. Gel permeation resolved three -protease variants in leaf extracts but only two in callus samples. Rechromatography of the large molecular weight fraction in a cation-exchange column produced three and two variants for leaf and callus, respectively. The present results suggest that there are at least five variants of sulfhydryl protease in tobacco leaf and three in callus tissue and that tobacco Fraction I protein can be metabolized by both leaf and callus proteases.

GENETIC DIVERSITY OF THE CYTOPLASM IN TRITICUM AND AEGILOPS. VIII. FRACTION I PROTEIN OF 39 CYTOPLASMS

ABSTRACT The electrophoretic characteristics of the cytoplasmically controlled large subunit of the Fraction I protein of 36 alloplasmic and three euplasmic control lines are reported. These lines, representing the cytoplasms of 32Triticum and Aegilops species, had either H- or L-type large subunits in their Fraction I protein; the diploid Triticum and most Aegilops species, including Ae. bicornisand Ae. sharonensis,had the L-type subunits; whereas, all the polyploid Triticum species (emmer, timopheevi, common wheats), Ae. speltoides, Ae. aucheri,and Ae. longissimahad H-type subunits. Therefore, section Sitopsis of Aegilops exhibits interspecific heterogeneity. The H-type is believed to have originated in the Sitopsis section from an L-type subunit because of the prevalence of the latter among the diploid species.