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United States Department of Agriculture

Agricultural Research Service

Title: Malate Synthesis in Alfalfa Root Nodules Is Controlled by Five Malate Dehydrogenase Genes

Authors
item Driscoll, Brian - UNIVERSITY OF MINNESOTA
item Miller, Susan - UNIVERSITY OF MINNESOTA
item Gregerson, Robert - UNIVERSITY OF MINNESOTA
item Gantt, J - UNIVERSITY OF MINNESOTA
item Vance, Carroll

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: July 18, 1996
Publication Date: N/A

Technical Abstract: Because of malate's multifunctional role in nodules, synthesis is controlled by numerous forms of malate dehydrogenase (MDH). To further understand how nodule malate metabolism is genetically regulated, we have isolated the cDNAs encoding alfalfa nodule MDH. Four separate MDH cDNA were isolated from an alfalfa root nodule library: mitochondrial-mMDH; glyoxysomal-gMDH; cytosolic-cMDH; and nodule enhanced-neMDH. The plastid- pMDH was isolated from an alfalfa leaf cDNA library. Complementation tests revealed that plasmids carrying gMDH, mMDH, cMDH, or neMDH rescued an E. coli Mdh**- mutant, while plasmids carrying pMDH did not. Analysis of various tissues showed that mMDH, cMDH, and gMDH are expressed relatively uniformly in all tissues, while pMDH expression occurred mainly in leaves and cotyledons. By contrast, neMDH mRNA was several-fold higher in nodules than any other tissue. The neMDH and cMDH enzyme was purified from complemented E. coli Mdh**- mutants and antibodies produced to the enzyme protein. Antibodies to the two proteins were immunologically distinct. The nodule enhanced nature of neMDH was also evident on protein blots, with nodules having 3- to 5-fold more neMDH protein than other organs. Amounts of neMDH protein were reduced some 60 to 70% in ineffective nodules. Initial kinetic analysis for malate saturation of activity indicate that cMDH has typical saturation kinetics, while neMDH has an unusual biphasic saturation curve and it is not saturated at concentrations of malate approaching 90 mM. The data support the hypothesis that nodules show high expression of a specific heretofore unidentified MDH and that this neMDH has unusual malate saturation kinetics that may be related to the high organic acid concentrations found in root nodules.

Last Modified: 7/30/2014
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