title: Detection of proteins related to starch synthase activity in the developing mungbean (Vigna radiata L.).
abstract: Proteins associated with starch synthase (SS) activities were identified in immature mungbeans (Vigna radiata L. cv KPS1). Seed soluble extract was separated by native-PAGE and subjected to in situ activity staining. The gel zymogram located starch-enzyme complex bands. The soluble extract was also partitioned by preparative-IEF and screened for SS activity using radioactive assay. IEF fractions eluted within pH 4-6 revealed enriched SS activity of 145-fold. Parallel comparison of the protein profiles among the activity stained enzyme complex and the active isoelectric focused fractions on SDS-PAGE depicted three SS-activity-related proteins with molecular size of 32, 53, and 85 kDa. The 85 kDa protein, however, was identified to be methionine synthase by MALDI-TOF analysis and should be a protein physically associated with the active SS. Polyclonal antibodies raised from eluted native enzyme complex neutralized up to 90% activity and antigenically recognize the other 53 and 32 kDa proteins on Western blot. Antibodies raised from the two individual denatured proteins were able to neutralize SS activities near 60% separately, indicating that the 53 kDa and 32 proteins associated with SS activity are potentially involved in starch biosynthesis during mungbean seed development.
authors.affiliation: Department of Food Science, Biotechnology Division, College of Life and Resource Sciences, National Taiwan Ocean University, 2 Pei-Ning Rd., Keelung, Taiwan 20224. irisko@mail.ntou.edu.tw
authors.forename: Yuan-Tih
authors.initials: YT
authors.lastname: Ko
authors.forename: Chun-Hsu
authors.initials: CH
authors.lastname: Pan
authors.forename: Ya-Ting
authors.initials: YT
authors.lastname: Lee
authors.forename: Jin-Yi
authors.initials: JY
authors.lastname: Chang
mesh_terms: D015153:Blotting, Western; D005591:Chemical Fractionation; D004591:Electrophoresis, Polyacrylamide Gel; D007887:Fabaceae / Q000201:enzymology*; D007106:Immune Sera / Q000494:pharmacology; D007525:Isoelectric Focusing; D010936:Plant Extracts / Q000737:chemistry; D012639:Seeds / Q000201:enzymology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013214:Starch Synthase / Q000032:analysis* / Q000276:immunology / Q000378:metabolism*
chemical_list: D007106:Immune Sera; D010936:Plant Extracts; D013214:Starch Synthase