Starch biosynthesis: further evidence against the primer nonreducing-end mechanism and evidence for the reducing-end two-site insertion mechanism.
Two reactions were studied with three varieties of starch granules from maize, wheat, and rice. In Reaction-I, the granules were reacted with 1 mM ADP-[(14)C]Glc and in Reaction-II, a portion of the granules from Reaction-I was reacted with 1 mM ADP-Glc. The starch granules were solubilized and reacted with the exo-acting glucoamylase and beta-amylase to an extent of 50% or less of the (14)C-label. The amounts of (14)C-labeled products from glucoamylase and beta-amylase were nearly equal for Reaction-I and Reaction-II. If the addition had been to the nonreducing ends of primers, Reaction-II would not have given any labeled products from the hydrolysis of glucoamylase and beta-amylase. These results indicate that the elongation of the starch chain is the addition of D-glucose to the reducing end by a de novo two-site insertion mechanism and not by the addition of D-glucose to the nonreducing end of a primer. This is in conformity with previous results in which starch granules were pulsed with ADP-[(14)C]Glc and chased with nonlabeled ADP-Glc, giving (14)C-labeled D-glucitol from the pulsed starch and a significant decrease in (14)C-labeled D-glucitol from the chased starch on reducing with NaBH(4) and hydrolyzing with glucoamylase [Carbohydr. Res.2002, 337, 1015-1022]. It also is in conformity with the inhibition of starch synthesis that occurs when putative primers are added to starch granule-ADP-Glc digests, indicating that the elongation is not by the nonreducing-end primer mechanism [Carbohydr. Res.2005, 340, 245-255].
Mukerjea
Rupendra
R
Laboratory of Carbohydrate Chemistry and Enzymology, Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University Ames, 50011, USA.
Robyt
John F
JF
D000245:Adenosine Diphosphate Glucose / Q000378:metabolism; D002250:Carbon Radioisotopes; D008320:Maltose / Q000378:metabolism; D008956:Models, Chemical; D012275:Oryza / Q000737:chemistry / Q000378:metabolism; D013213:Starch / Q000096:biosynthesis* / Q000737:chemistry; D014908:Triticum / Q000737:chemistry / Q000378:metabolism; D003313:Zea mays / Q000737:chemistry / Q000378:metabolism; D001614:beta-Amylase / Q000378:metabolism
D002250:Carbon Radioisotopes; D000245:Adenosine Diphosphate Glucose; D008320:Maltose; D013213:Starch; D001614:beta-Amylase