Amino acid transport : the special case of a H/L-glutamate cotransport system in Asparagus sprengeri mesophyll cells /

The addition of L-Glutamate (L-GLU) and L-Hethionine ~ulfoximine (L-HSO) to mechanically isolated. photosynthetically competent, Asparagus sprengeri mesophyll cells ~u~pended in 1mM CaS04 cau~ed an immediate transient alkalinization of the cell su~pension medium in both the light and dark. The alkalinization response was specific and stereospecific as none of the L-isomers of the other 19 protein amino acids tested or D-GLU gave this response. Uptake of 14C-L-GLU was stimulated by the light. The addition of non-radioactive L-GLU. or L-GLU analogs together with 14C-L-GLU showed that only L-GLU and L-HSO stimulated alkalinization whilst inhibiting the uptake of 14C-L-GLU. Both the L-GLU dependent alkalinization and the upt~ke of 14C-L-GLU were stimulated when the external pH was decreased from 6.5 to 5.5. Increasing external K+ concentrations inhibited the uptake of 14C-L-GLU. Fusicoccin (FC) stimulated uptake. The L-GLU dependent alkalinization re~ponse exhibited monophasic saturation kinetics while the uptake of 14C-L-GLU exhibited biphasic saturation kinetics. In addition to a saturable component. the uptake kinetics also showed a linear component of uptake. Addition of L-GLU and L-MSO caused internal acidification of the cell as measured by a change in the distribution of 14C-DMO. There was no change in K+ efflux when L-GLU was added. A H+ to L-GLUinflux stoichiometry of 3:1 wa~ mea~ured at an external I.-GLU concentration of O.5mM and increased with increasing external 13 L-QLU concentration. Metabolism of L-GLU was detected manometrlcally by observing an increase in COa evolution upon the addition of L-QLU and by detection of i*C02 evolution upon the addition of »*C-L-GLU. »*C02 evolution was higher in the dark than in the light. The data are consistent with the operation of a H+/L-QLO cotransport system. The data also show that attempts to quantify the stoichlometry of the process were complicated by the metabolism of L-GLU.

Rapid [Gamma]-amino acid synthesis and the inhibition of the growth and development of oblique banded leaf-roller larvae

The hypothesis that rapid y-aminobutyric acid (GABA) accumulation is a plant defense against phytophagous insects was investigated. Simulation of mechanical damage resulting from phytophagous insect activity increased soybean (Glycine max L.) leaf GABA 10- to 25-fold within 1 to 4 min. Pulverizing leaf tissue resulted in a value of 2. 15 (±O. 11 SE) ~mol GABA per gram fresh weight. Increasing the GABA levels in a synthetic diet from 1.6 to 2.6 Jlffiol GABA per gram fresh weight reduced the growth rates, developmental rates, total biomass (50% reduction), and survival rates (30% reduction) of cultured Oblique banded leaf-roller (OBLR) (Choristonellra rosacealla Harris) larvae. In field experiments OBLR larvae were found predominantly on young terminal leaves which have a reduced capacity to produce GABA in response to mechanical damage. Glutamate decarboxylase (GAD) is a cytosolic enzyme which catalyses the decarboxylation of L-Glu to GABA. GAD is a calmodulin binding enzyme whose activity is stimulated dramatically by increased cytosolic H+ or Ca2 + ion concentrations. Phytophagous insect activity will disrupt the cellular compartmentation of H+ and Ca2 +, activate GAD and subsequent GABA accumulation. In animals GABA is a major inhibitory neurotransmitter. The possible mechanisms resulting in GABA inhibited growth and development of insects are discussed.

Synthesis and utilization of guanidine catalysts for applications directed towards the preparation of butenolide and modified amino acid derivatives

Development of guanidine catalysts is explored through direct iminium chloride and amine coupling, alongside a 2-chloro-l,3-dimethyl-IH-imidazol-:-3-ium chloride (DMC) induced thiourea cyclization. Synthesized achiral catalyst N-(5Hdibenzo[ d,t][1,3]diazepin-6(7H)-ylidene)-3,5-bis(trifluoromethyl) aniline proved unsuccessful towards O-acyl migrations, however successfully catalyzed the vinylogous aldol reaction between dicbloro furanone and benzaldehyde. Incorporating chirality into the guanidine catalyst utilizing a (R)-phenylalaninol auxiliary, generating (R)-2-((5Hdibenzo[ d,t] [1,3 ]diazepin-6(7H)-ylidene ) amino )-3 -phenylpropan-l-ol, demonstrated enantioselectivity for a variety of adducts. Highest enantiomeric excess (ee) was afforded between dibromofuranone and p-chlorobenzaldehyde, affording the syn conformation in 96% ee and the anti in 54% ee, with an overall yield of30%. Attempts to increase asymmetric induction were focused on incorporation of axial chirality to the (R)phenylalaninol catalyst using binaphthyl diamine. Incorporation of (S)-binaphthyl exhibited destructive selectivity, whereas incorporation of (R)-binaphthyl demonstrated no effects on enantioselectivity. Current studies are being directed towards identifying the catalytic properties of asymmetric induction with further studies are being aimed towards increasing enantioselectivity by increasing backbone steric bulk.