CHARACTERIZATION OF ADPGLUCOSE PYROPHOSPHORYLASE FROM A STARCH-DEFICIENT MUTANT OF ARABIDOPSIS-THALIANA (L)

被引:31
作者
LI, L [1 ]
PREISS, J [1 ]
机构
[1] MICHIGAN STATE UNIV,DEPT BIOCHEM,E LANSING,MI 48824
基金
美国国家科学基金会;
关键词
D O I
10.1016/0008-6215(92)85074-A
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
A starch-deficient mutant of Arabidopsis thaliana (L.) has only approximately 5% of ADPglucose pyrophosphorylase activity but accumulates approximately 40% of starch as the wild-type. ADPglucose pyrophosphorylase from the mutant and wild-type leaves was partially purified and characterized. The activity of each enzyme was inhibited by an antibody raised against purified spinach-leaf ADPglucose pyrophosphorylase. When analyzed by Western-blot hybridization after SDS-PAGE, the partially purified mutant enzyme was deficient in the larger of the two subunits observed in the wild-type enzyme. However, by Western-blot hybridization after native PAGE, the mutant enzyme demonstrated two equally stained cross-reactive bands instead of a single band for the wild-type enzyme. Of the mutant enzyme activity, > 95% was found in the lower band. The mass of the native enzyme from either wild-type or mutant was approximately 210 kD, as determined by gel filtration, and those of the subunits were approximately 54 and 48 kD for the wild-type enzyme and 48 kD for the mutant enzyme. The mutant ADPglucose phyrophosphorlase, just as the wild-type enzyme, was activated allosterically by 3-P-glycerate and inhibited by P(i). However, the mutant enzyme required higher concentrations of 3-P-glycerate for maximal activation and was more sensitive to inhibition by P(i) than the wild-type enzyme. In the presence of saturating concentrations of 3-P-glycerate, the K(m) values for the mutant enzyme for ATP, Glc-1-P, and Mg2+ were approximately 6-, approximately 5-, and approximately 2-fold higher, respectively, than those of the wild-type enzyme. Changes in the kinetics of the mutant enzyme may be due to a deficiency of one of the two subunits, and/or modification of the remaining subunit.
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页码:227 / 239
页数:13
相关论文
共 17 条
  • [1] IDENTIFICATION AND MOLECULAR CHARACTERIZATION OF SHRUNKEN-2 CDNA CLONES OF MAIZE
    BHAVE, MR
    LAWRENCE, S
    BARTON, C
    HANNAH, LC
    [J]. PLANT CELL, 1990, 2 (06) : 581 - 588
  • [2] BURNETTE WN, 1981, ANAL BIOCHEM, V112, P2195
  • [3] PURIFICATION OF SPINACH LEAF ADP-GLUCOSE PYROPHOSPHORYLASE
    COPELAND, L
    PREISS, J
    [J]. PLANT PHYSIOLOGY, 1981, 68 (05) : 996 - 1001
  • [4] GHOSH HP, 1966, J BIOL CHEM, V241, P4491
  • [5] SULFONYLUREA-RESISTANT MUTANTS OF ARABIDOPSIS-THALIANA
    HAUGHN, GW
    SOMERVILLE, C
    [J]. MOLECULAR & GENERAL GENETICS, 1986, 204 (03): : 430 - 434
  • [6] ADPGLUCOSE PYROPHOSPHORYLASE IS ENCODED BY DIFFERENT MESSENGER-RNA TRANSCRIPTS IN LEAF AND ENDOSPERM OF CEREALS
    KRISHNAN, HB
    REEVES, CD
    OKITA, TW
    [J]. PLANT PHYSIOLOGY, 1986, 81 (02) : 642 - 645
  • [7] ISOLATION AND CHARACTERIZATION OF A STARCHLESS MUTANT OF ARABIDOPSIS-THALIANA (L) HEYNH LACKING ADPGLUCOSE PYROPHOSPHORYLASE ACTIVITY
    LIN, TP
    CASPAR, T
    SOMERVILLE, C
    PREISS, J
    [J]. PLANT PHYSIOLOGY, 1988, 86 (04) : 1131 - 1135
  • [8] A STARCH DEFICIENT MUTANT OF ARABIDOPSIS-THALIANA WITH LOW ADPGLUCOSE PYROPHOSPHORYLASE ACTIVITY LACKS ONE OF THE 2 SUBUNITS OF THE ENZYME
    LIN, TP
    CASPAR, T
    SOMERVILLE, CR
    PREISS, J
    [J]. PLANT PHYSIOLOGY, 1988, 88 (04) : 1175 - 1181
  • [9] SUBUNIT STRUCTURE OF SPINACH LEAF ADPGLUCOSE PYROPHOSPHORYLASE
    MORELL, MK
    BLOOM, M
    KNOWLES, V
    PREISS, J
    [J]. PLANT PHYSIOLOGY, 1987, 85 (01) : 182 - 187
  • [10] OBERFELDER R, 1989, FOCUS, V11, P1