Nitrogen (N) is an essential requirement for kernel growth in maize (Zea mays); however, little is known about how N assimilates are metabolized in young earshoots during seed development. The objective of this study was to assess amino acid metabolism in cob and spikelet tissues during the critical 2 weeks following silking. Two maize hybrids were grown in the field for 2 years at two levels of supplemental N fertilizer (0 and 168 kg N/ha). The effects of the reproductive sink on cob N metabolism were examined by comparing pollinated to unpollinated earshoots. Earshoots were sampled at 2, 8, 14, and 18 d after silking; dissected into cob, spikelet, and/or pedicel and kernel fractions; then analyzed for amino acid profiles and key enzyme activities associated with amino acid metabolism. Major amino acids in the cob were glutamine (Gln), aspartic acid (Asp), asparagine (Asn), glutamate, and alanine. Gln concentrations dropped dramatically from 2 to 14 d after silking in both pollinated and unpollinated cobs, whereas all other measured amino acids accumulated over time in unpollinated spikelets and cobs, especially Asn. N supply had a variable effect on individual amino acid levels in young cobs and spikelets, with Asn being the most notably enhanced. We found that the cob performs significant enzymatic interconversions among Gln, alanine, Asp, and Asn during early reproductive development, which may precondition the N assimilate supply for sustained kernel growth. The measured amino acid profiles and enzymatic activities suggest that the Asn to Gln ratio in cobs may be part of a signal transduction pathway involving aspartate aminotransferase, Gln synthetase, and Asn synthetase to indicate plant N status for kernel development.Though there are numerous studies concerning growth and development of maize (Zea mays) grain, there is a general lack of knowledge concerning the physiology of the cob tissues, to which kernels are attached. This is despite the fact that the cob tissues are the link between vegetative source and reproductive sink tissues. In addition, approximately one-half of the earshoot (minus husk and shank) is cob material at silking, with the cob proportion decreasing as the grain develops. Early studies on the physical characteristics of the earshoot and cob go back more than a century (Harshberger, 1893). More recent research has examined cob shape (Srinivas et al., 1991;Bhattacharya et al., 1996;Orr et al., 1997;Pearson, 2000) and the genetic control of cob color (Nguetta and Cross, 1997;Sidorenko et al., 1999). A few studies have also focused on sugars (BeMiller and Hoffmann, 1972), nitrogen (N;Crawford et al., 1982), minerals (Mozafar, 1990), and antioxidants (Cevallos and Cisneros, 2003) contained within maize earshoots.A typical view of maize cobs is that they serve as a temporary storage organ and as a conveyor of nutrients to the developing kernels (Crawford et al., 1982). A larger metabolic role of the cob in kernel growth, however, can be inferred from the observation that an attached cob ...

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The carboxyterminal processing protease of D1 protein (CtpA) is predicted to be an excellent target for a general broad-spectrum herbicide. The gene for spinach CtpA has been expressed in Escherichia coli. The expressed protein that was found mainly in inclusion bodies has been purified and refolded on a nickel-chelate column. Active recombinant CtpA was recovered. Two assays for CtpA activity were developed, a medium-throughput HPLC assay using a fluorescent substrate and a high-throughput assay based on fluorescence polarization capable of application in a high-throughput 96-well plate format. This high-throughput assay was developed to screen chemistry for CtpA inhibitors. Native spinach CtpA was partially purified and the native and recombinant enzymes were compared kinetically for their K(m) and V(max) values using different peptide substrates. Native CtpA partially purified from spinach was shown to have similar kinetic properties to recombinant CtpA. Antibodies developed against the recombinant protein were used to estimate the in planta abundance of the native enzyme in spinach. Since only a small proportion of the recombinant protein is refolded during isolation and it appears that only a small proportion of this enzyme is active, size-exclusion chromatography and light scattering experiments were performed on rCtpA in order to gain insight into its structure and the reasons why most of the protein is not active. The use of rCtpA to screen for herbicidal compounds and the more general question of how good a herbicide target the enzyme is are discussed.

~,bstract Initiation factor elF2 binds GTP and promotes the binding of methionyl-tRNA to ribosomes. Biochemical and sequence evidence suggests that the GTP might bind to either the /3-or 3,-subunit of elF2. Mutations were made in the NKXD consensus elements found in both subunits and individual mutant forms were overexpressed in transiently transfected COS-1 cells. the effect on the translational efficiency of a reporter mRNA for dihydrofolate reductase was monitored. Mutations in the y-subunit cause severe repression of protein synthesis, whereas those in the/]-subunit are only mildly inhibitory. The results support the view that GTP binds exclusively to the 3,-subunit.