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UBC Theses and Dissertations

Plant protein chemotaxonomy : I. Disc Electrophoresis of Lasthenia seed albumins and globulins : II. Partial characterization and sequence studies of Sambucus ferredoxin Altosaar, Illimar

Abstract

Proteins indirectly reflect, via messenger-RNA, the information coded in DNA, and are thus considered to be tertiary semantides. Since proteins are amenable to comparative analyses, they can provide a chemical basis for a molecular phylogeny. This dissertation reports the application of two approaches to the study of plant relationships using protein characters. The first was an electrophoretic comparison of seed storage proteins from all twenty taxa of the genus Lasthenia. Albumin and globulin fractions were extracted from dormant achenes. Each protein sample was fractionated by disc electrophoresis in basic 7% polyacrylamide gels. Mean R[sub P] values, coefficients of variation, and 95% confidence intervals were calculated for both types of protein bands. Similarity coefficients, calculated from the distribution of homologous bands, were used to produce dendrograms. Affinities among the taxa differ from the conventional taxonomy of the genus. The second approach involved the purification and characterization of an iron-sulphur protein from a higher plant and comparison with ferredoxin from other species. Ferredoxin was isolated from leaves of Sambucus raaemosa L. by the following procedure: 1) homogenization in buffered 50% acetone-water, 2) ion-exchange chromatography on several columns of DEAE-cellulose, and 3) finally purified in good yield by gel filtration. The UV and visible spectrum showed maxima at 277, 331, 423, and 466 nm. The protein sustained an initial photoreduction rate of 86 μmoles NADP per mg chlorophyll per hour. The amino acid composition was found to be Lys₅, His₂, Arg₁, Asx₁₁, Thr₅, Ser₇, Glx₁₇, Pro₆ , Gly₇, Ala₆₋₇, Cys₄, Val₈, He₅, Leu₇, Tyr₃, Phe₂ , and Trp₁. The molecule has a molecular weight of 10,700 and contains 2 atoms of iron. The amino-terminal sequence is Ala-Thr and the carboxyl-terminal sequence is Leu-Thr-Ala. These properties are discussed in relation to those of other angiosperm ferredoxins. Experiments were performed to investigate the feasibility of sequencing this ferredoxin.

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