Photodegradable antimicrobial agents: towards structure optimization.

Antibiotic resistance continues to be an ominous threat facing human health globally and urgent action is required to limit the loss of human life. The pollution of antibiotics into the environment is one of the drivers behind the crisis. With this in mind, we have developed novel photodecomposable antimicrobial agents based on an ethanolamine scaffold, which upon photoirradiation decomposes into two major inactive fragments. Herein we describe our further work on the synthesis of novel ethanolamines with a particular focus on structure activity relationship, resulting in four new active compounds which photodecomposed into inactive fragments.

Identify Primary Air Pollution Sources of BTEX by Using Positive Matrix Factorization (PMF): A Case Study of Ho Chi Minh City, Vietnam.

Ho Chi Minh City (HCMC) is one of the main socioeconomic and financial centers of Vietnam. The city also faces serious air pollution. However, the city polluted with benzene, toluene, ethylbenzene, and xylene (BTEX) has rarely been studied. We used positive matrix factorization (PMF) to analyze BTEX concentrations measured at two sampling locations to identify the main sources of BTEX in HCMC. The locations represented residential area (i.e., To Hien Thanh) and industrial area (i.e., Tan Binh Industrial Park). At the To Hien Thanh location, the average concentrations of benzene, ethylbenzene, toluene, and xylene were 6.9, 14.4, 4.9, and 12.7 µg/m3, respectively. At the Tan Binh location, the average concentrations of benzene, ethylbenzene, toluene, and xylene were 9.8, 22.6, 2.4, and 9.2 µg/m3, respectively. The results showed that PMF was a reliable model for source apportionment in HCMC. Traffic activities were the main sources of BTEX. Besides, industrial activities also contributed to BTEX emissions, especially the location near the industrial park. The majority of BTEXs at the To Hien Thanh sampling site come from traffic sources accounting for 56.2%. Activities from traffic and photochemical reactions (42.7%) and industrial sources (40.5%) were the main sources affecting BTEX emissions at the sampling site of Tan Binh Industrial Park. This study can be used as a reference for mitigation solutions to reduce the BTEX emission in HCMC.

Essential Oils from the Leaves, Stem, and Roots of Blumea lanceolaria (Roxb.) Druce in Vietnam: Determination of Chemical Composition, and In Vitro, In Vivo, and In Silico Studies on Anti-Inflammatory Activity.

Blumea lanceolaria (Roxb.) Druce, a flowering plant, is used for treating cancer and inflammatory diseases. In this study, we determined the chemical composition of the EOs extracted from the leaves (LBEO), stem (SBEO), and roots (RBEO) of B. lanceolaria and analyzed their anti-inflammation potential. Overall, 30 compounds representing 99.12%, 98.44%, and 96.89% of total EO constituents of the leaves, stem, and roots, respectively, were identified using GC-MS. ELISA, Western blotting, and qRT-PCR studies showed that LBEO, SBEO, and RBEO inhibited multiple steps in the inflammatory responses in the RAW 264.7 cell model, including NO production; TNF-α, IL-6, iNOS, and COX-2 transcription and translation; and phosphorylation of IκBα and p65 of the NF-κB pathway. In the carrageenan-induced paw edema model, all three EOs inhibited paw edema at both early and delayed phases. Molecular docking studies indicated that the main components of B. lanceolaria EOs (BEOs) targeted and inhibited major components of inflammation-related pathways, including the arachidonic acid metabolic pathway, NF-κB pathway, and MAPK pathway. We present the first study to characterize the chemical composition of BEOs and confirm their potent anti-inflammatory effects in in vitro, in vivo, and in silico analysis. These results can facilitate the development of effective anti-inflammatory drugs with limited side effects in the future.

Cultivation of Caenorhabditis elegans on new cheap monoxenic media without peptone.

The study of species biodiversity within the Caenorhabditis genus of nematodes would be facilitated by the isolation of as many species as possible. So far, over 50 species have been found, usually associated with decaying vegetation or soil samples, with many from Africa, South America and Southeast Asia. Scientists based in these regions can contribute to Caenorhabditis sampling and their proximity would allow intensive sampling, which would be useful for understanding the natural history of these species. However, severely limited research budgets are often a constraint for these local scientists. In this study, we aimed to find a more economical, alternative growth media to rear Caenorhabditis and related species. We tested 25 media permutations using cheaper substitutes for the reagents found in the standard nematode growth media (NGM) and found three media combinations that performed comparably to NGM with respect to the reproduction and longevity of C. elegans. These new media should facilitate the isolation and characterization of Caenorhabditis and other free-living nematodes for the researchers in the poorer regions such as Africa, South America, and Southeast Asia where nematode diversity appears high.

Molecular typing of conjunctivitis-causing adenoviruses in Hanoi, Vietnam from 2017 to 2019 and complete genome analysis of the most prevalent type (HAdV-8).

Adenoviral conjunctivitis is a common epidemic worldwide. In Vietnam, up to 80,000 patients are infected with adenoviral conjunctivitis annually. However, there are few investigations on the pathogenic adenoviruses that cause conjunctivitis. In total, 120 eye-swab samples were collected from patients with viral conjunctivitis symptoms in Hanoi, Vietnam from 2017 to 2019. Human adenoviruse (HAdV) was detected in 67 samples (55.83%) using polymerase chain reaction amplification of at least one of three HAdV-specific marker genes (hexon, penton, and fiber). Of the 67 HAdV samples, 46 samples could be analyzed by all three marker genes. DNA sequence analysis and phylogenetic tree building based on the three marker genes from the 46 HAdV samples revealed five different HAdV types associated with conjunctivitis in Hanoi, including HAdV-3 (4.3%), HAdV-4 (2.2%), HAdV-8 (89.1%), HAdV-37 (2.2%), and a potential recombinant type between types HAdV-8 and HAdV-3 (2.2%). This showed that HAdV-8 was the most common type identified in Hanoi. Complete genome analysis of HAdV-8 isolated from a Vietnamese patient (VN2017) using Sanger sequencing revealed 34 unique nucleotide changes, indicating that the adenovirus continuously accumulates new mutations. Hence, continuous surveillance of HAdV-8 changes in Vietnam is necessary in the future.

Improved genomic resources for the black tiger prawn (Penaeus monodon).

World production of farmed crustaceans was 7.8 million tons in 2016. While only making up approximately 10% of world aquaculture production, crustaceans are generally high-value species and can earn significant export income for producing countries. Viet Nam is a major seafood producing country earning USD 7.3 billion in 2016 in export income with shrimp as a major commodity. However, there is a general lack of genomic resources available for shrimp species, which is challenging to obtain due to the need to deal with large repetitive genomes, which characterize many decapod crustaceans. The first tiger prawn (P. monodon) genome assembly was assembled in 2016 using the standard Illumina PCR-based pair-end reads and a computationally-efficient but relatively suboptimal assembler, SOAPdenovo v2. As a result, the current P. monodon draft genome is highly fragmented (> 2 million scaffolds with N50 length of <1000 bp), exhibiting only moderate genome completeness (< 35% BUSCO complete single-copy genes). We sought to improve upon the recently published P. monodon genome assembly and completeness by generating Illumina PCR-free pair-end sequencing reads to eliminate genomic gaps associated with PCR-bias and performing de novo assembly using the updated MaSurCA de novo assembler. Furthermore, we scaffolded the assembly with low coverage Nanopore long reads and several recently published deep Illumina transcriptome paired-end sequencing data, producing a final genome assembly of 1.6 Gbp (1,211,364 scaffolds; N50 length of 1982 bp) with an Arthropod BUSCO completeness of 96.8%. Compared to the previously published P. monodon genome assembly from China (NCBI Accession Code: NIUS01), this represents an almost 20% increase in the overall BUSCO genome completeness that now consists of more than 90% of Arthropod BUSCO single-copy genes. The revised P. monodon genome assembly (NCBI Accession Code: VIGR01) will be a valuable resource to support ongoing functional genomics and molecular-based breeding studies in Vietnam.

Homologous Lympho-Epithelial Kazal-type Inhibitor Domains Delay Blood Coagulation by Inhibiting Factor X and XI with Differential Specificity.

Despite being initially identified in the blood filtrate, LEKTI is a 15-domain Kazal-type inhibitor mostly known in the regulation of skin desquamation. In the current study, screening of serine proteases in blood coagulation cascade showed that LEKTI domain 4 has inhibitory activity toward only FXIa, whereas LEKTI domain 6 inhibits both FXIa and FXaB (bovine FXa). Nuclear magnetic resonance structural and dynamic experiments plus molecular dynamics simulation revealed that LEKTI domain 4 has enhanced backbone flexibility at the reactive-site loop. A model of the LEKTI-protease complex revealed that FXaB has a narrower S4 pocket compared with FXIa and hence prefers only small side-chain residues at the P4 position, such as Ala in LEKTI domain 6. Mutational studies combined with a molecular complex model suggest that both a more flexible reactive-site loop and a bulky residue at the P4 position make LEKTI domain 4 a weaker but highly selective inhibitor of FXIa.

Structural basis for the bacterial membrane insertion of dermcidin peptide, DCD-1L.

Human dermcidin (DCD) is an antimicrobial peptide secreted constitutively by sweat glands. The anionic derivative, DCD-1L, comprises of the N-terminal 47 residues of DCD and one additional leucine residue. A previous NMR structure of DCD-1L in 50% TFE showed a partial helical conformation, and its crystal structure in the presence of Zn2+ outlined a hexameric linear α-helical bundle. Three different models to describe membrane insertion were proposed but no conclusion was drawn. In the current study, the NMR structure of DCD-1L in SDS micelles showed an "L-shaped" molecule with three fully formed α-helices connected by flexible turns. Formation of these helices in DCD-1L in the presence of POPG vesicles suggests that the acidic C-terminal region of DCD-1L can suppress the binding of DCD-1L to POPG vesicles at basic but not acidic pH. Mutation of charged residues on the N-terminal and C-terminal regions of DCD-1L cause differences in POPG binding, suggesting distinct functional roles for these two regions. Charged residues from these two regions are also found to differentially affect Zn2+ coordination and aggregation of DCD-1L in the absence or presence of SDS, as monitored by 1D NMR. Our data agrees with one of the three models proposed.

Structure of AcrH-AopB Chaperone-Translocator Complex Reveals a Role for Membrane Hairpins in Type III Secretion System Translocon Assembly.

Type III secretion systems (T3SSs) are adopted by pathogenic bacteria for the transport of effector proteins into host cells through the translocon pore composed of major and minor translocator proteins. Both translocators require a dedicated chaperone for solubility. Despite tremendous efforts in the past, structural information regarding the chaperone-translocator complex and the topology of the translocon pore have remained elusive. Here, we report the crystal structure of the major translocator, AopB, from Aeromonas hydrophila AH-1 in complex with its chaperone, AcrH. Overall, the structure revealed unique interactions between the various interfaces of AopB and AcrH, with the N-terminal "molecular anchor" of AopB crossing into the "N-terminal arm" of AcrH. AopB adopts a novel fold, and its transmembrane regions form two pairs of helical hairpins. From these structural studies and associated cellular assays, we deduced the topology of the assembled T3SS translocon; both termini remain extracellular after membrane insertion.

Specificity and inhibitory mechanism of andrographolide and its analogues as antiasthma agents on NF-κB p50.

Andrographolide (1) is a diterpenoid lactone with an α,ß-unsaturated lactone group that inhibits NF-κB DNA binding. Andrographolide reacts with the nucleophilic Cys62 of NF-κB p50 through a Michael addition at the Δ(12(13)) exocylic double bond to form a covalent adduct. Using computer docking, site-directed mutagenesis, and mass spectrometry, the noncovalent interactions between andrographolide and additional binding site residues other than Cys62 were found to be essential for the covalent incorporation of andrographolide. Furthermore, the addition reaction of andrographolide on Cys62 was highly dependent on the redox conditions and on the vicinity of nearby, positively charged Arg residues in the conserved RxxRxR motif. The reaction mechanisms of several of the analogues were determined, showing that 14-deoxy-11,12-didehydroandrographolide (8) reacts with NF-κB p50 via a novel mechanism distinct from andrographolide. The noncovalent interaction and redox environment of the binding site should be considered, in addition to the electrophilicity, when designing a covalent drug. Analogues similar in structure appear to use distinct reaction mechanisms and may have very different cytotoxicities, e.g., compound 6.

A quantitative chemical proteomics approach to profile the specific cellular targets of andrographolide, a promising anticancer agent that suppresses tumor metastasis.

Drug target identification is a critical step toward understanding the mechanism of action of a drug, which can help one improve the drug's current therapeutic regime and expand the drug's therapeutic potential. However, current in vitro affinity-chromatography-based and in vivo activity-based protein profiling approaches generally face difficulties in discriminating specific drug targets from nonspecific ones. Here we describe a novel approach combining isobaric tags for relative and absolute quantitation with clickable activity-based protein profiling to specifically and comprehensively identify the protein targets of andrographolide (Andro), a natural product with known anti-inflammation and anti-cancer effects, in live cancer cells. We identified a spectrum of specific targets of Andro, which furthered our understanding of the mechanism of action of the drug. Our findings, validated through cell migration and invasion assays, showed that Andro has a potential novel application as a tumor metastasis inhibitor. Moreover, we have unveiled the target binding mechanism of Andro with a combination of drug analog synthesis, protein engineering, and mass-spectrometry-based approaches and determined the drug-binding sites of two protein targets, NF-κB and actin.