Initiation of polysaccharide synthesis in plants
Date of Award
Doctor of Philosophy (Ph.D.)
Biochemistry and Molecular Biology
First Committee Member
William J. Whelan, Committee Chair
Mammalian glycogen synthesis is initiated on a protein primer, termed glucogenin, which remains covalently bound to the mature macromolecule. Glycogenin (Mr 38kDa) is an autocatalytic enzyme that undergoes self-glucosylation to form a maltooctaose unit attached to tyrosine 194 on itself. The carbohydrate unit so formed is then converted into the glycogen macromolecule by glycogen synthesizing enzymes.A new protein has now been discovered in the plant kingdom that has properties similar to glycogenin. This protein, isolated from sweet corn, has a molecular weight of 42kDa and, like glycogenin, is also a divalent cation dependent autoglucosylating enzyme. Furthermore, the sweet corn self-glucosylating protein (SGP) cross reacted with a polyclonal antibody to glycogenin, implying that there is structural similarity between the two proteins. Nine tryptic peptides were isolated and sequenced from the sweet corn SGP (which together comprised approximately 50% of the protein's structure), including the peptide containing the glucosylation site. The amino acid sequences of these peptides are unique and have no sequence homology either to glycogenin or to any other protein or peptide sequences in the protein data banks.Amino acid and sequence analyses on the glucosylated tryptic peptide isolated from sweet corn SGP after undergoing self-glucosylation identified arginine as the amino acid to which the glucose was attached. This was confirmed by ion-spray mass spectrometry. Furthermore, tandem mass spectrometry indicated that only one glucose residue was linked to the arginine which was hydrolyzable by beta-glucosidase. In undergoing autoglucosylation, therefore, the sweet corn SGP added a single glucose unit to an arginine residue on itself joined through a beta-linkage. This arginine-beta-glucose bond in the sweet corn SGP represents a novel protein-glucose linkage.Similar to glycogenin, the plant enzyme also appears to prime carbohydrate synthesis. It can be converted into a higher molecular weight molecule in a cell free extract with nucleotide sugars. During development, the protein appears in advance of the onset of starch production and reaches its peak concentration together with the starch synthesizing enzymes. This is consistent with the protein being a precursor of starch.
Biology, Molecular; Chemistry, Biochemistry
Gieowar Singh, Dave, "Initiation of polysaccharide synthesis in plants" (1994). Dissertations from ProQuest. 3233.