title: In vitro stabilization and minimum active component of polygalacturonic acid synthase involved in pectin biosynthesis.
abstract: Polygalacturonic acid (PGA) synthase successively transfers galacturonic acid to oligogalacturonic acid by an alpha1,4-linkage to synthesize PGA, the backbone of plant pectic homogalacturonan. PGA synthase has not been purified to date due to its instability in vitro. In this study, we found stable conditions in vitro and separated a minimum active component of the enzymes from pea and azuki bean epicotyls. The PGA synthase lost its activity in 500 mM of sodium chloride or potassium chloride, while it was relatively stable at low salt concentrations. Under low salt concentrations, three peaks bearing PGA synthase activity were separated, by gel filtration and sucrose density gradient centrifugation. The molecular masses of these enzymes solubilized with 3-[(3-cholamidopropyl)dimethyl-ammonio]propanesulfonic acid were estimated to be 21,000, 5,000, and 590 kDa. The two higher molecular mass PGA synthases converted to smaller PGA synthase proteins when treated with high salt concentrations, while retaining their activity, indicating that PGA synthase has a minimum active component for its activity.
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, Osaka 560-0043, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Ohashi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Ishimizu
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Akita
authors.forename: Sumihiro
authors.initials: S
authors.lastname: Hase
mesh_terms: D002021:Buffers; D002793:Cholic Acids / Q000737:chemistry; D002850:Chromatography, Gel; D004789:Enzyme Activation; D007887:Fabaceae / Q000201:enzymology; D007700:Kinetics; D010368:Pectins / Q000096:biosynthesis* / Q000378:metabolism*; D011485:Protein Binding; D012995:Solubility