Designing antimicrobial peptides using deep learning and molecular dynamic simulations.

With the emergence of multidrug-resistant bacteria, antimicrobial peptides (AMPs) offer promising options for replacing traditional antibiotics to treat bacterial infections, but discovering and designing AMPs using traditional methods is a time-consuming and costly process. Deep learning has been applied to the de novo design of AMPs and address AMP classification with high efficiency. In this study, several natural language processing models were combined to design and identify AMPs, i.e. sequence generative adversarial nets, bidirectional encoder representations from transformers and multilayer perceptron. Then, six candidate AMPs were screened by AlphaFold2 structure prediction and molecular dynamic simulations. These peptides show low homology with known AMPs and belong to a novel class of AMPs. After initial bioactivity testing, one of the peptides, A-222, showed inhibition against gram-positive and gram-negative bacteria. The structural analysis of this novel peptide A-222 obtained by nuclear magnetic resonance confirmed the presence of an alpha-helix, which was consistent with the results predicted by AlphaFold2. We then performed a structure-activity relationship study to design a new series of peptide analogs and found that the activities of these analogs could be increased by 4-8-fold against Stenotrophomonas maltophilia WH 006 and Pseudomonas aeruginosa PAO1. Overall, deep learning shows great potential in accelerating the discovery of novel AMPs and holds promise as an important tool for developing novel AMPs.

Single-Disulfide Conopeptide Czon1107, an Allosteric Antagonist of the Human α3ß4 Nicotinic Acetylcholine Receptor.

Conopeptides are peptides in the venom of marine cone snails that are used for capturing prey or as a defense against predators. A new cysteine-poor conopeptide, Czon1107, has exhibited non-competitive inhibition with an undefined allosteric mechanism in the human (h) α3ß4 nicotinic acetylcholine receptors (nAChRs). In this study, the binding mode of Czon1107 to hα3ß4 nAChR was investigated using molecular dynamics simulations coupled with mutagenesis studies of the peptide and electrophysiology studies on heterologous hα3ß4 nAChRs. Overall, this study clarifies the structure-activity relationship of Czon1107 and hα3ß4 nAChR and provides an important experimental and theoretical basis for the development of new peptide drugs.

Determination of urine oxalate level in rats with renal calcium oxalate calculus by high-performance liquid chromatography.

OBJECTIVE: To establish a method of high-performance liquid chromatography (HPLC) for determining the urine oxalate levle in rats with renal calcium oxalate calculus. METHODS: Totally 24 SPF Wistar healthy male rats were randomly divided into control group(n=12)and ethylene glycol (EG) group (n=12). Rats in EG group were administered intragastrically with 2% ammonium chloride (AC)2 ml/rat per day+1% ethylene glycol (EG), along with free access to drinking water.The control group was fed with deionized water, along with the intragastric administration of normal saline (1 ml per day). Twenty-eight days after modelling, the 24-hour urine samples were collected, and the urine oxalic acid levels were determined using HPLC and the results were compared with those of catalytic spectrophotometry using oxidation of methyl. During the HPLC, the samples were separated on Aglient 5TC-C18 (250×4.6 mm,5 Μm), eluted with mixture of methanol (0.1 mol/L) and ammonium acetate (15:85) at 1.2 ml/min, and detected at 314 nm, with the column temperature being 20 ℃. RESULTS: The standard curves of high and low concentrations of oxalic acid were y=5909.1x+378730, R² =0.9984 and y=7810.5x-16635, R² =0.9967,respectively. The lowest detectable concentration in this method was 5 Μg/ml. The linear high concentration range of oxalate stood at 62.50-2000.00 Μg/ml, and the linear low concentration range of oxalate stood at 6.25-100.00 Μg/ml. Its average recovery was 95.1%, and its within-day and day-to-day precisions were 3.4%-10.8% and 3.8%-9.4%. Both HPLC and catalytic spectrophotometry showed significantly higher urinary oxalic acid concentration and 24 h urine oxalate level in EG group compared with the control group [urinary oxalic acid concentration: (736.35 ± 254.52) Μg/ml vs.(51.56 ± 36.34) Μg/ml,(687.35 ± 234.53) Μg/ml vs.(50.24 ± 42.34) Μg/ml;24 h urine oxalate level: (11.23 ± 4.12)mg vs.(0.87 ± 0.45)mg,(9.89 ± 3.55)mg vs. (0.77 ± 0.65)mg; all P<0.01]. No statistically significant difference was observed in the results of urinary oxalate concentration and 24 h urine oxalate level between HPLC and potassium chromate oxidation of methyl red spectrophotometry (all P>0.05). CONCLUSION: HPLC is a simple, rapid, and precise method in detecting urine oxalate level in rats with renal calcium oxalate calculus, with high recovery rate.

Stewed Rhubarb Decoction Ameliorates Adenine-Induced Chronic Renal Failure in Mice by Regulating Gut Microbiota Dysbiosis.

This study aimed to investigate the protective effect of Stewed Rhubarb (SR) decoction on chronic renal failure (CRF) through the regulation of gut microbiota. Using a CRF mouse model induced by a 0.2% adenine diet, we proved that SR decoction (2.0 g crude SR/kg) significantly reduced the levels of urea and creatinine in plasma of CRF mice, accompanied by the improvement of renal fibrosis and tubular atrophy, amelioration of inflammation, and inhibition of aquaporins damage. Also, SR decoction alleviated gut barrier damage, indicative of the elevated mRNA expression of intestinal mucins and tight junctions. By 16S rDNA sequencing, SR decoction reshaped the imbalanced gut microbiota in CRF mice by statistically reversing the abundance changes of a wide range of intestinal bacteria at family and genus levels, which further led to balance in the production of intestinal metabolites, including short-chain fatty acids (acetic acid, propionic acid, and valeric acid), indole, and bile acids (TUDCA and CDCA). Inversely, SR decoction failed to repress the occurrence of CRF in mice with gut microbiota depletion, confirming the essential role of gut microbiota in SR decoction-initiated protection against CRF. In summary, SR decoction can improve adenine-induced CRF in mice by remolding the structure of destructed gut microbiota community. Our findings shed light on the clinical application of SR decoction in nephropathy treatment.