ASPARTATE-AMINOTRANSFERASE IN ALFALFA ROOT-NODULES .2. IMMUNOLOGICAL DISTINCTION BETWEEN 2 FORMS OF THE ENZYME

被引:33
作者
FARNHAM, MW [1 ]
MILLER, SS [1 ]
GRIFFITH, SM [1 ]
VANCE, CP [1 ]
机构
[1] UNIV MINNESOTA,DEPT AGRON & PLANT GENET,ST PAUL,MN 55455
关键词
D O I
10.1104/pp.93.2.603
中图分类号
Q94 [植物学];
学科分类号
071001 ;
摘要
Aspartate aminotransferase (AAT), a key enzyme in the biosynthesis of aspartate and asparagine, occurs as two forms in alfalfa (Medicago sativa L.), AAT-1 and AAT-2. Both forms were purified to near homogeneity, and high titer polyclonal antibodies produced to the native proteins. Alfalfa AAT-1 was purified from root suspension culture cells, while AAT-2 was purified from effective root nodules. Antibodies prepared to AAT-1 and used as probes for western blots readily recognized native and SDS forms of AAT-1 but did not recognize either native or SDS forms of AAT-2. Conversely, antibodies to AAT-2 readily recognized native and SDS forms of AAT-2 but did not recognize AAT-1. Immunotitrations further confirmed the immunological distinction between AAT-1 and AAT-2. AAT-1 antibodies immunotitrated 100% of the in vitro activity of purified AAT-1 but had no effect on AAT-2 in vitro activity. Likewise, AAT-2 antibodies removed 100% of the in vitro activity of purified AAT-2 but did not affect AAT-1 in vitro activity. Sequential titration of total AAT activity from roots and nodules showed that AAT-1 comprised the major form (62%) of AAT in roots, while AAT-2 was the predominant form (90%) in nodules. Last, SDS-PAGE western blots showed that the molecular masses of AAT-1 and AAT-2 were 42 and 40 kilodaltons, respectively. These data indicate that AAT is under the control of at least two distinct genes in alfalfa.
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页码:603 / 610
页数:8
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