1. Co-localization of glutamate and tubulin in putative excitatory neurons of the hippocampus and amygdala: an immunohistochemical study using monoclonal antibodies
J E Madl, A J Beitz, A A Larson, R Kuriyama, A J McDonald, C Sellitto Neuroscience . 1989;30(2):405-21. doi: 10.1016/0306-4522(89)90261-3.
Tubulin and glutamate were immunohistochemically localized in the hippocampus and amygdala of rats using monoclonal antibodies to gamma-Glu-Glu (Glu-1) and glutaraldehyde-fixed glutamate (Glu-2), respectively. Glu-2 was shown to be selectively immunoreactive for glutaraldehyde-fixed Glu using enzyme-linked immunoassays and inhibition enzyme-linked immunoassays. Glu-1 was immunoreactive with tubulin on immunoblots of brain homogenates. However, only tubulin with a glutamate carboxy-terminal appeared to be immunoreactive with Glu-1 since tubulin from Chinese hamster ovary cells was not immunoreactive on immunoblots unless the tubulin was first treated with carboxypeptidase. Intense immunocytochemical staining by Glu-1 of hippocampus and amygdala was co-localized in the same neurons as the immunocytochemical staining for glutaraldehyde-fixed Glu produced by Glu-2. The distribution of immunostaining in the brain by Glu-1 was very similar to the distribution of immunostaining produced by Glu-2. The major difference was that glutamate-like immunoreactivity, visualized by Glu-2 staining, was intense in the nuclei of neurons, while nuclei were unstained by Glu-1. The distribution of immunostaining by these monoclonal antibodies was very similar to that reported in previous studies using other antibodies to Glu. All granule cells in the area dentata of the hippocampus exhibited intense immunoreactivity with both antibodies. Immunoreactivity was also observed in the stratum lucidum of CA3, the zone of termination of mossy fiber axons of granule cells. The majority of pyramidal cells in CA1, and many pyramidal cells in CA3 of the hippocampus were immunoreactive. In addition, it appeared that all of the pyramidal cells in the subiculum exhibited immunoreactivity. Light, diffuse immunoreactivity was observed in the neuropil of the hippocampus and subiculum. Most perikarya in the amygdala were characterized by light to moderate Glu-1 immunoreactivity and moderate to intense Glu-2 immunoreactivity. Fairly intense Glu-1 and Glu-2 immunoreactivity was seen in some neurons of the lateral nucleus, basolateral nucleus, lateral subdivision of the central nucleus, and the periamygdaloid cortex. The morphology of immunostained neurons in the lateral and basolateral nuclei indicates that the majority of these cells correspond to the pyramidal class 1 neurons described in previous Golgi studies.(ABSTRACT TRUNCATED AT 400 WORDS)
2. Comparative study on the novel umami-active peptides of the whole soybeans and the defatted soybeans fermented soy sauce
Chun Cui, Xiping Zhu, Dongxiao Sun-Waterhouse, Wei Wang, Jiahui Chen J Sci Food Agric . 2021 Jan 15;101(1):158-166. doi: 10.1002/jsfa.10626.
Background:Two kinds of soy sauce produced via fermentation of the whole soybeans and the defatted soybeans (soy sauce termed 'SSS' and 'SSD', respectively) were subjected to the treatment using aqueous ethanol solutions with high concentrations. Then tasty peptides were separated from SSS and SSD by sensory guided fractionation, using macroporous resin and reverse-phase high-performance liquid chromatography (RP-HPLC), and identified by ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UPLC-Q-TOF-MS/MS).Results:The result showed that umami-active fractions and ≤ 3 kDa peptides were mainly concentrated in the supernatants resulted from the treatment with 60% ethanol and the precipitates resulted from the treatment with 80% ethanol. The contents of ammonia nitrogen, non-salt solids, bitter amino acids, amino acids in peptides and the proportion of ≤ 3 kDa peptides in SSS were higher than those in SSD. Sixteen and fourteen tasty peptides were separated from SSS and SSD, among them five dipeptides (γ-Glu-Glu, Glu-Glu, γ-Glu-Cys, γ-Glu-Leu, Glu-Leu and Ile-Glu) with a glutamic acid residue were identified both in SSS and SSD, which have been reported as umami/kokumi-active peptides in soy sauce. Several peptides identified from SSS (Thr-Gly-Cys, Gly-Leu-Glu, Val-Glu-Ala-Leu and Gly-Gly-Gly-Glu) and SSD (Asp-Arg, Asp-Ala-Glu, Glu-Val-Cys and Gly-Gly-Gly-Glu) are tasty and/or umami-active peptides but have not been reported as tasty peptides in the past.Conclusion:Most of the peptides separated from SSD and SSS could impart an umami-enhancing effect on soy sauce, and the marginally more kokumi and bitterness hydrophobic peptides were found in SSS than SSD. © 2020 Society of Chemical Industry.
3. Feasibility of synthesizing γ-[Glu](n≥1)-Gln using high solid concentrations and glutaminase from Bacillus amyloliquefaciens as the catalyst
Chun Cui, Dongxiao Sun-Waterhouse, Wei Wang, Jian Xiong, Runmei Tang Food Chem . 2020 Apr 25;310:125920. doi: 10.1016/j.foodchem.2019.125920.
Effects of using high solid concentrations on the synthesis of γ-[Glu](n≥1)-Gln catalyzed by glutaminase from Bacillus amyloliquefaciens were examined in this study. An increment in solid concentration from 10% to 50% (w/w) increased the extent of synthesis of γ-[Glu](n≥1)-Gln, based on the analyses of amino acid composition and amino nitrogen content. Size-exclusion high-performance liquid chromatography analysis revealed an increase in molecular mass of γ-[Glu](n≥1)-Gln resulting from the increase of solid concentration from 10% to 50% (w/w). UPLC-Q-TOF-MS/MS analysis showed that the enzymatic reaction mixtures post γ-glutamyl transpeptidation contained γ-Glu-Gln, γ-Glu-Glu-Gln, γ-Glu-Glu-Glu-Gln, γ-Glu-Glu-Glu-Glu-Gln and γ-Glu-Glu-Glu-Glu-Glu-Gln. The intensity of each γ-[Glu](n=1,2,3,4,5)-Gln produced at a solid concentration of 50% (w/w) was higher than that at 10% (w/w). These findings indicated the potential of such an energy- and water-efficient approach for synthesizing γ-[Glu](n≥1)-Gln at high solid concentrations.
