Occurrence of D-amino acids in natural products

Since the identified standard genetic code contains 61 triplet codons of three bases for the 20 L-proteinogenic amino acids(AAs),no D-AA should be found in natural products.This is not what is observed in the living world.D-AAs are found in numerous natural compounds produced by bacteria,algae,fungi,or marine animals,and even vertebrates.A review of the literature indicated the existence of at least 132 peptide natural compounds in which D-AAs are an essential part of their structure.All compounds are listed,numbered and described herein.The two biosynthetic routes leading to the presence of D-AA in natural products are:non-ribosomal peptide synthesis(NRPS),and ribosomally synthesized and post-translationally modified peptide(RiPP)synthesis which are described.The methods used to identify the AA chirality within naturally occurring peptides are briefly discussed.The biological activity of an all-L synthetic peptide is most often completely different from that of the D-containing natural compounds.Analyzing the selected natural compounds showed that D-Ala,D-Val,D-Leu and D-Ser are the most commonly encountered D-AAs closely followed by the non-proteinogenic D-allo-Thr.D-Lys and D-Met were the least prevalent D-AAs in natu-rally occurring compounds.

Promising role of D-amino acids in irritable bowel syndrome

Irritable bowel syndrome(IBS)is an important health care concern.Alterations in the microbiota of the gut-brain axis may be linked to the pathophysiology of IBS.Some dietary intake could contribute to produce various metabolites including Damino acids by the fermentation by the gut microbiota.D-amino acids are the enantiomeric counterparts of L-amino acids,in general,which could play key roles in cellular physiological processes against various oxidative stresses.Therefore,the presence of D-amino acids has been shown to be linked to the protection of several organs in the body.In particular,the gut microbiota could play significant roles in the stability of emotion via the action of D-amino acids.Here,we would like to shed light on the roles of D-amino acids,which could be used for the treatment of IBS.

Site-specific incorporation of reduction-controlled guest amino acids into proteins for cucurbituril recognition

Protein recognition using host-guest recognition approach is of great interest but has been limited mainly to the protein N-terminal residues.Here,we site-specific incorporated two novel non-canonical amino acids containing supramolecular guest motifs into protein via an expanded genetic code.Through Staudinger reduction reactions,the encoded unnatural residues on protein becoming activated and can be specifically recognized by cucurbit[7]uril(CB[7])and cucurbit[8]uril(CB[8]).We demonstrated that enzyme containing guest amino acid incorporated near the active site can be reversibly regulated by CB[7]recognition,and CB[8]recognition induces protein dimerization.These amino acids will make useful addition to the supramolecular toolbox for protein targeting using molecular recognition approaches.

Metal-organic framework hybrid materials of ZIF-8/RGO for immobilization of D-amino acid dehydrogenase

Immobilization of D-amino acid dehydrogenase(DAADH)by the assembly of peptide linker was studied for the biosynthesis of Dphenylalanine.Hybrid material of zeolitic imidazolate framework-8(ZIF-8)combined with reduced graphene oxide(RGO)was applied for the immobilization of DAADH from Ureibacillus thermosphaericus.The recovery rate of DAADH/ZIF-8/RGO was 165.6%.DAADH/ZIF-8/RGO remained 53.4%of its initial activity at 50°C for 10 h while the free enzyme was inactivated.DAADH/ZIF-8/RGO maintained 70.5%activity in hyperalkaline solution with pH 12.Kinetic parameters indicated that DAADH/ZIF-8/RGO had greater affinity of phenylpyruvate as V_(max)/K_(m)of DAADH/ZIF-8/RGO was 1.27-fold than free enzyme.After seven recycles,the activity of DAADH/ZIF-8/RGO remained 64.3%.Furthermore,one-step separation and in situ immobilization of DAADH by ZIF-8/RGO/Ni was carried out with 1.5-fold activity enhancement.Combining peptide linker and metal-organic framework(MOF)immobilization,thermostability and activity of the immobilized DAADH were significantly improved.

Microbial D-amino acids and marine carbon storage

In nature, there are two conformational types of amino acids: L- and D-isomers. The L-amino acids are the predominant form and are used mainly for protein synthesis, while the D-amino acids are few in quantity but more diverse in terms of their biological functions. D-amino acids are produced by many marine microbes, which are important players in carbon and energy cycles in the ocean. As the major constituent of the marine organic carbon pool, D-amino acids can persist in the water column for a long time before being further transformed by chemical or biological processes or transported through physical processes(such as absorption and aggregation). This article reviews the microbial synthesis of D-amino acids, their physiological function and metabolism in microbes, and the contribution of D-amino acids as a carbon source to the oceanic carbon reservoir.

Colorimetric detection of D-amino acids based on anti-aggregation of gold nanoparticles

A new method has been proposed to realize the visual detection of D-amino acids （DAAs） via the antiaggregation of 4-mercaptobenzoic acid modified gold nanoparticles （AuNPs） in the presence of D-amino acid oxidase （DAAO）. The negatively charged AuNPs were prepared using sodium citrate as a reducer and stabilizer. The presence of 4-mercaptobenzoic acid （4-MBA） and Cu2＋ induces the aggregation of AuNPs, resulting in a color change from ruby red to royal purple. However, DAAO could oxidize DAAs to generate H2O2. In the presence of H2O2, the mercapto （-SH） group in 4-mercaptobenzoic acid can be oxidized to form a disulfide （-S-S-） bond. Based on these facts, the pre-incubation of DAAs and 4-mercaptobenzoic acid with DAAO would significantly reduce the concentration of free 4-mercaptobenzoic acid molecules, thus the aggregation of AuNPs was interrupted since due to the lack of inducer. As the concentration of DAAs increases, the color of the AuNPs solution would progress from royal purple to ruby red. Consequently, DAAs could be monitored by the colorimetric response of AuNPs using a UV-vis spectrophotometer or even naked eyes. This DAAO mediated visual detection method could determine D- alanine （D-Ala） as a representative DAA with concentrations ranging from 1.5 × 10^-7mol L 1 to 3.0 × 10^-5 mol L^-1, and the detection limit was as low as 7.5 × 10^-8 mol L^-1. The proposed method is convenient, low-cost and free of complex equipment, making it feasible to analyze the concentration of D-AIa in real samples of β-amyloid peptide （Aβ1-42）.

D-Tyr-tRNA^(Tyr) Deacylase,a New Role in Alzheimer's-associated Disease in SAMP8 Mice

Objective To assess the expression level of D-Tyr-tRNATyr deacylase(DTD) in SAMP8 mice and speculate the function of DTD in disorders associated with Alzheimer's disease(AD).Methods Altogether 12 SAMP8 mice and 12 SAMR1 mice were used in this study.Semi-quantita-tive reverse transcription-polymerase chain reaction(RT-PCR) and Western blot were performed to detect the mRNA and protein levels of DTD in the mice.Purified DTD protein was injected into lateral ventricle to investigate the function of DTD in SAMP mice.The behavior of the mice was tested by using a Step-through Test System.Results Both mRNA and protein levels of DTD were found to be significantly lower in SAMP8 mice compared with those in SAMR1 mice(P<0.05).In vivo injection of DTD protein did not lead to an obvious change in behavior of SAM mice.Conclusions DTD might function in the process of AD-associated pathology and could possibly participate in physiology process in a long-term manner to orchestrate with other regulators in order to maintain the balance of organism.

Efficacy of probiotics on the modulation of gut microbiota in the treatment of diabetic nephropathy

Diabetic nephropathy(DN)is a major cause of end-stage renal disease,and therapeutic options for preventing its progression are insufficient.The number of patients with DN has been increasing in Asian countries because of westernization of dietary lifestyle,which may be associated with the following changes in gut microbiota.Alterations in the gut microbiota composition can lead to an imbalanced gastrointestinal environment that promotes abnormal production of metabolites and/or inflammatory status.Functional microenvironments of the gut could be changed in the different stages of DN.In particular,altered levels of short chain fatty acids,D-amino acids,and reactive oxygen species biosynthesis in the gut have been shown to be relevant to the pathogenesis of the DN.So far,evidence suggests that the gut microbiota may play a key role in determining networks in the development of DN.Interventions directing the gut microbiota deserve further investigation as a new protective therapy in DN.In this review,we discuss the potential roles of the gut microbiota and future perspectives in the protection and/or treatment of kidneys.

Enzymatic cascade systems for D-amino acid synthesis:progress and perspectives

D-amino acids,different from the ubiquitous L-amino acids,are recognized as the“unnatural”amino acids.The applications of D-amino acids have drawn increasing interest from researchers in recent years,and D-amino acids are widely used in various industries,including for food products,pharmaceuticals,and agricultural chemicals.Inspired by the prevalent appli-cations,many synthetic methods for D-amino acids have been developed,which are mainly divided into chemical synthetic methods and biosynthetic methods.Chemical synthesis of D-amino acids has a variety of disadvantages such as multiple reaction steps,low yields,low reaction rates,and difficulties in product extraction.Thus,biosynthetic methods utilizing enzymes are attracting increasing attention because they are more energy-saving and environmentally friendly compared to traditional chemical synthesis.Among all enzymatic methods,multi-enzymatic cascade catalytic methods have significant advantages,such as lower costs,no need for intermediate separation,and higher catalytic efficiency,which is ascribed to the spatial proximity of biocatalysts.In this review,advances in multi-enzyme cascade catalytic systems as well as chemo-enzymatic approaches to synthesize D-amino acids are discussed.

A dual-function selection system enables positive selection of multigene CRISPR mutants and negative selection of Cas9-free progeny in Arabidopsis

The CRISPR/Cas9 technology revolutionizes targeted gene knockout in diverse organisms including plants.However,screening edited alleles,particularly those with multiplex editing,from herbicide-or antibiotic-resistant transgenic plants and segregating out the Cas9 transgene represent two laborious processes.Current solutions to facilitate these processes rely on different selection markers.Here,by taking advantage of the opposite functions of a D-amino acid oxidase(DAO)in detoxifying D-serine and in metabolizing non-toxic D-valine to a cytotoxic product,we develop a DAO-based selection system that simultaneously enables the enrichment of multigene edited alleles and elimination of Cas9-containing progeny in Arabidopsis thaliana.Among five DAOs tested in Escherichia coli,the one encoded by Trigonopsis variabilis(TvDAO)could confer slightly stronger D-serine resistance than other homologs.Transgenic expression of TvDAO in Arabidopsis allowed a clear distinction between transgenic and nontransgenic plants in both D-serine-conditioned positive selection and D-valine-conditioned negative selection.As a proof of concept,we combined CRISPR-induced single-strand annealing repair of a dead TvDAO with D-serine-based positive selection to help identify transgenic plants with multiplex editing,where D-serine-resistant plants exhibited considerably higher co-editing frequencies at three endogenous target genes than those selected by hygromycin.Subsequently,D-valine-based negative selection successfully removed Cas9 and TvDAO transgenes from the survival offspring carrying inherited mutations.Collectively,this work provides a novel strategy to ease CRISPR mutant identification and Cas9 transgene elimination using a single selection marker,which promises more efficient and simplified multiplex CRISPR editing in plants.

Effects of d-Phenylalanine as a biocide enhancer of THPS against the microbiologically influenced corrosion of C1018 carbon steel

Microbiologically influenced corrosion(MIC) is caused by biofilms such as those of sulfate reducing bacteria(SRB). To mitigate MIC, biocide treatment is often needed. Tetrakis(hydroxymethyl) phosphonium sulfate(THPS) is an environmentally friendly biocide that is often used in the oil and gas industry. However, its prolonged use leads to biocide resistance, leading to dosage escalation. A biocide enhancer can be used to slow down the trend. In recent years, d-amino acids have been investigated as an enhancer for THPS and other biocides. Published works used anaerobic vials and flow devices, which could not reveal the real-time changes of the biocide treatment on corrosion. In this work, it was proven that the biocide enhancement effects of d-Phenylalanine(d-Phe) on THPS against the Desulfovibrio vulgaris biofilm on C1018 carbon steels could be assessed in real time using linear polarization resistance and electrochemical impedance spectroscopy to collaborate sessile cell count, weight loss and pitting depth data. The results showed that 500 ppm(w/w) d-Phe effectively enhanced 80 ppm THPS against MIC by the D. vulgaris(a corrosive SRB) biofilm. The sessile cell count and pit depth were all reduced with the enhancement of d-Phe.

Chemical modifications of proteins and their applications in metalloenzyme studies

Protein chemical modifications are important tools for elucidating chemical and biological functions of proteins.Several strategies have been developed to implement these modifications,including enzymatic tailoring reactions,unnatural amino acid incorporation using the expanded genetic codes,and recognition-driven transformations.These technologies have been applied in metalloenzyme studies,specifically in dissecting their mechanisms,improving their enzymatic activities,and creating artificial enzymes with non-natural activities.Herein,we summarize some of the recent efforts in these areas with an emphasis on a few metalloenzyme case studies.