His-Glu

Background: This study has elucidated moisture dynamics in the soybean peptide, Ser-His-Glu-Cys-Asn (SHECN) powder by using dynamic vapor sorption (DVS) and nuclear magnetic resonance (NMR). We also tried to investigate the effects of moisture absorption on the biological activity and chemical properties of SHECN with some effective methods such as mid-infrared (MIR) spectroscopy and gas chromatography-mass spectrometry (GC-MS). Results: DVS results showed that the moisture absorption of SHECN could reach a maximum of 33%, and the SHECN powder after synthesis actually existed in a trihydrate state of SHECN.3H2 O. Low-field NMR revealed that three water proportions including strong combined water, binding water and bulk water were involved in SHECN moisture absorption and absored water dominantly existed in the form of combined water. Magnetic resonance imaging (MRI) and MIR spectroscopy results indicated that moisture absorption could change the morphology and structure of SHECN. After moisture absorption at 50% and 75% relative humidity, 19 volatiles were identified by GC-MS analysis. Additionally, this study showed that a part of reductive groups in SHECN was oxidized and its antioxidant ability declined significantly (P < 0.05) after moisture absorption. Conclusion: Water absorbed into SHECN powder can significantly change its microstructure and cause its activity to decrease. We must prevent SHECN from absorbing moisture during storage because the water can accelerate the oxidation of samples and promote microbial reactions. © 2016 Society of Chemical Industry.

The catalytic mechanism of Pdx2 was studied with atomic detail employing the computational ONIOM hybrid QM/MM methodology. Pdx2 employs a Cys-His-Glu catalytic triad to deaminate glutamine to glutamate and ammonia - the source of the nitrogen of pyridoxal 5'-phosphate (PLP). This enzyme is, therefore, a rate-limiting step in the PLP biosynthetic pathway of Malaria and Tuberculosis pathogens that rely on this mechanism to obtain PLP. For this reason, Pdx2 is considered a novel and promising drug target to treat these diseases. The results obtained show that the catalytic mechanism of Pdx2 occurs in six steps that can be divided into four stages: (i) activation of Cys87 , (ii) deamination of glutamine with the formation of the glutamyl-thioester intermediate, (iii) hydrolysis of the formed intermediate, and (iv) enzymatic turnover. The kinetic data available in the literature (19.1-19.5 kcal mol-1 ) agree very well with the calculated free energy barrier of the hydrolytic step (18.2 kcal.mol-11 ), which is the rate-limiting step of the catalytic process when substrate is readily available in the active site. This catalytic mechanism differs from other known amidases in three main points: i) it requires the activation of the nucleophile Cys87 to a thiolate; ii) the hydrolysis occurs in a single step and therefore does not require the formation of a second tetrahedral reaction intermediate, as it is proposed, and iii) Glu198 does not have a direct role in the catalytic process. Together, these results can be used for the synthesis of new transition state analogue inhibitors capable of inhibiting Pdx2 and impair diseases like Malaria and Tuberculosis.

Background: Using high-performance liquid chromatography/tandem mass spectrometry, two novel antioxidant pentapeptides [Ser-His-Glu-Cys-Asn (SHECN) and Leu-Pro-Phe-Ala-Met (LPFAM)] were identified from 1-3-kDa soybean protein hydrolysates (SPH). The MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay was used to evaluate cytotoxicity in HepG2 cells. Antioxidant activity was measured using in vitro assays, including the cellular antioxidant activity assay (CAA), 2,2-diphenyl-1-picrylhydrazyl or 2,2-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) inhibition, and oxygen radical absorbance capacity (ORAC) assays. Finally, the secondary structure was determined using circular dichroism (CD). Results: The results revealed that two novel peptides were nontoxic and possessed antioxidant activity. SHECN had significantly higher antioxidant activity than LPFAM (P < 0.05). The CAA value of SHECN was 776.22 µmol QE 100 g-1 . SHECN also showed significant DPPH inhibition (70.18 ± 4.06%) and ABTS inhibition (88.16 ± 0.76%). It had normalized ORAC values of 0.3000 ± 0.0070 µmol GE mg-1 and 0.0900 ± 0.0020 µmol TE mg-1 , respectively. The results of the CD analysis demonstrated that, compared to LPFAM, which had much lower antioxidant activity, SHECN had a high β-sheet content and reduced α-helix content. Conclusion: The results indicated that SHECN possessed high antioxidant activity. A higher β-sheet content and lower content levels of α-helix appear to be correlated with antioxidant activity. © 2016 Society of Chemical Industry.