Mechanistic and structural insights into EstS1 esterase: A potent broad-spectrum phthalate diester degrading enzyme.

Phthalate diesters are important pollutants and act as endocrine disruptors. While certain bacterial esterases have been identified for phthalate diesters degradation to monoesters, their structural and mechanistic characteristics remain largely unexplored. Here, we highlight the potential of the thermostable and pH-tolerant EstS1 esterase from Sulfobacillus acidophilus DSM10332 to degrade high molecular weight bis(2-ethylhexyl) phthalate (DEHP) by combining biophysical and biochemical approaches along with high-resolution EstS1 crystal structures of the apo form and with bound substrates, products, and their analogs to elucidate its mechanism.

Expression, purification, kinetics, and crystallization of non-heme mononuclear iron enzymes: Biphenyl, Phthalate, and Terephthalate dioxygenases.

Non-heme iron oxygenases constitute a versatile enzyme family that is crucial for incorporating molecular oxygen into diverse biomolecules. Despite their importance, only a limited number of these enzymes have been structurally and functionally characterized. Surprisingly, there remains a significant gap in understanding how these enzymes utilize a typical architecture and reaction mechanism to catalyze a wide range of reactions.

Structural and functional characterization of the nucleotide-binding domains of ABCA4 and their role in Stargardt disease.

ABCA4 is an ATP-binding cassette (ABC) transporter that prevents the buildup of toxic retinoid compounds by facilitating the transport of N-retinylidene-phosphatidylethanolamine across membranes of rod and cone photoreceptor cells. Over 1500 missense mutations in ABCA4, many in the nucleotide-binding domains (NBDs), have been genetically linked to Stargardt disease. Here, we show by cryo-EM that ABCA4 is converted from an open outward conformation to a closed conformation upon the binding of adenylyl-imidodiphosphate.

Sulisobenzone is a potent inhibitor of the global transcription factor Cra.

Transcription is carried out by the RNA polymerase and is regulated through a series of interactions with transcription factors. Catabolite activator repressor (Cra), a LacI family transcription factor regulates the virulence gene expression in Enterohaemorrhagic Escherichia coli (EHEC) and thus is a promising drug target for the discovery of antivirulence molecules. Here, we report the crystal structure of the effector molecule binding domain of Cra from E.

Crystal structure and molecular characterization of NADP-farnesol dehydrogenase from cotton bollworm, Helicoverpaarmigera.

Farnesol dehydrogenase (FDL) orchestrates the oxidation reaction catalyzing farnesol to farnesal, a key step in the juvenile hormone (JH) biosynthesis pathway of insects and hence, represents a lucrative target for developing insect growth regulators (IGRs). However, information on the structural and functional characterization of JH-specific farnesol dehydrogenase in insects remains elusive. Herein, we identified a transcript that encodes farnesol dehydrogenase (HaFDL) from Helicoverpa armigera, a major pest of cotton.

Conformational flexibility enables catalysis of phthalate cis-4,5-dihydrodiol dehydrogenase.

Phthalate cis-4,5-dihydrodiol dehydrogenase (PhtC), the second enzyme of the phthalate catabolic pathway, catalyzes the dehydrogenation of cis-4,5-dihydrodiol phthalate (DDP). Here, we report the structural and biochemical characterization of PhtC from Comamonas testosteroni KF1 (PhtC). With biochemical experiments, we have determined the enzyme's catalytic efficiency (k/K) with DDP as 2.

Structural Insights into Dihydroxylation of Terephthalate, a Product of Polyethylene Terephthalate Degradation.

Biodegradation of terephthalate (TPA) is a highly desired catabolic process for the bacterial utilization of this polyethylene terephthalate (PET) depolymerization product, but to date, the structure of terephthalate dioxygenase (TPDO), a Rieske oxygenase (RO) that catalyzes the dihydroxylation of TPA to a -diol, is unavailable. In this study, we characterized the steady-state kinetics and first crystal structure of TPDO from Comamonas testosteroni KF1 (TPDO). TPDO exhibited substrate specificity for TPA (/ = 57 ± 9 mM s).

Molecular insights into substrate recognition and catalysis by phthalate dioxygenase from Comamonas testosteroni.

Phthalate, a plasticizer, endocrine disruptor, and potential carcinogen, is degraded by a variety of bacteria. This degradation is initiated by phthalate dioxygenase (PDO), a Rieske oxygenase (RO) that catalyzes the dihydroxylation of phthalate to a dihydrodiol. PDO has long served as a model for understanding ROs despite a lack of structural data.

Deciphering the enigma of missing DNA binding domain of LacI family transcription factors.

Catabolite repressor activator (Cra) is a member of the LacI family transcriptional regulator distributed across a wide range of bacteria and regulates the carbon metabolism and virulence gene expression. In numerous studies to crystallize the apo form of the LacI family transcription factor, the N-terminal domain (NTD), which functions as a DNA-binding domain, has been enigmatically missing from the final resolved structures. It was speculated that the NTD is disordered or unstable and gets cleaved during crystallization.

MD simulation and MM/PBSA identifies phytochemicals as bifunctional inhibitors of SARS-CoV-2.

The global spread of SARS-CoV-2 has resulted in millions of fatalities worldwide, making it crucial to identify potent antiviral therapeutics to combat this virus. We employed structure-assisted virtual screening to identify phytochemicals that can target the two proteases which are essential for SARS-CoV-2 replication and transcription, the main protease and papain-like protease. Using virtual screening and molecular dynamics, we discovered new phytochemicals with inhibitory activity against the two proteases.

Biophysical and In-Silico Studies of Phytochemicals Targeting Chorismate Synthase from Drug-Resistant Moraxella Catarrhalis.

Chorismate serves as a crucial precursor for the synthesis of many aromatic compounds essential for the survival and virulence in various bacteria and protozoans. Chorismate synthase, a vital enzyme in the shikimate pathway, is responsible for the formation of chorismate from enolpyruvylshikimate-3-phosphate (EPSP). Moraxella catarrhalis is reported to be resistant to many beta-lactam antibiotics and causes chronic ailments such as otitis media, sinusitis, laryngitis, and bronchopulmonary infections.

Characterization of dye-decolorizing peroxidase from Bacillus subtilis.

The dye-decolorizing peroxidases (DyPs) belong to a unique heme peroxidase family for their biotechnological potential to detoxify synthetic dyes. In this work, we have biochemically and structurally characterized the dye-decolorizing peroxidase from Bacillus subtilis (BsDyP). The biochemical studies of BsDyP demonstrate that pH 4.

Engineering Burkholderia xenovorans LB400 BphA through Site-Directed Mutagenesis at Position 283.

Biphenyl dioxygenase (BPDO), which is a Rieske-type oxygenase (RO), catalyzes the initial dioxygenation of biphenyl and some polychlorinated biphenyls (PCBs). In order to enhance the degradation ability of BPDO in terms of a broader substrate range, the BphAE, BphAE, and BphAE variants were created from the parent enzymes BphAE, BphAE, and BphAE, respectively, by a substitution at one residue, Ser283Met. The results of steady-state kinetic parameters show that for biphenyl, the / values of BphAE, BphAE, and BphAE were significantly increased compared to those of their parent enzymes.

Biodegradation of phthalic acid esters (PAEs) and in silico structural characterization of mono-2-ethylhexyl phthalate (MEHP) hydrolase on the basis of close structural homolog.

Three bacterial strains capable of degrading phthalates namely Pseudomonas sp. PKDM2, Pseudomonas sp. PKDE1 and Pseudomonas sp.