ZP2 cleavage blocks polyspermy by modulating the architecture of the egg coat.

Following the fertilization of an egg by a single sperm, the egg coat or zona pellucida (ZP) hardens and polyspermy is irreversibly blocked. These events are associated with the cleavage of the N-terminal region (NTR) of glycoprotein ZP2, a major subunit of ZP filaments. ZP2 processing is thought to inactivate sperm binding to the ZP, but its molecular consequences and connection with ZP hardening are unknown.

Structure of the decoy module of human glycoprotein 2 and uromodulin and its interaction with bacterial adhesin FimH.

Glycoprotein 2 (GP2) and uromodulin (UMOD) filaments protect against gastrointestinal and urinary tract infections by acting as decoys for bacterial fimbrial lectin FimH. By combining AlphaFold2 predictions with X-ray crystallography and cryo-EM, we show that these proteins contain a bipartite decoy module whose new fold presents the high-mannose glycan recognized by FimH. The structure rationalizes UMOD mutations associated with kidney diseases and visualizes a key epitope implicated in cast nephropathy.

Cryo-EM structure of native human uromodulin, a zona pellucida module polymer.

Assembly of extracellular filaments and matrices mediating fundamental biological processes such as morphogenesis, hearing, fertilization, and antibacterial defense is driven by a ubiquitous polymerization module known as zona pellucida (ZP) "domain". Despite the conservation of this element from hydra to humans, no detailed information is available on the filamentous conformation of any ZP module protein. Here, we report a cryo-electron microscopy study of uromodulin (UMOD)/Tamm-Horsfall protein, the most abundant protein in human urine and an archetypal ZP module-containing molecule, in its mature homopolymeric state.

Crystal structure of ErmE - 23S rRNA methyltransferase in macrolide resistance.

Pathogens often receive antibiotic resistance genes through horizontal gene transfer from bacteria that produce natural antibiotics. ErmE is a methyltransferase (MTase) from Saccharopolyspora erythraea that dimethylates A2058 in 23S rRNA using S-adenosyl methionine (SAM) as methyl donor, protecting the ribosomes from macrolide binding. To gain insights into the mechanism of macrolide resistance, the crystal structure of ErmE was determined to 1.

Biosynthesis of leukotriene B.

Leukotriene B (LTB) is a lipid mediator derived from arachidonic acid (AA) by the sequential action of 5-lipoxygenase (5-LOX), 5-lipoxygenase-activating protein (FLAP) and LTA hydrolase (LTAH). It was initially recognized for its involvement in the recruitment of neutrophils and is one of the most potent chemotactic agents known to date. A large body of data has indicated that LTB plays a significant role in many chronic inflammatory diseases, such as arthritis, chronic obstructive pulmonary disease (COPD), cardiovascular disease, cancer and more recently, metabolic disorder.

Capturing LTA hydrolase in action: Insights to the chemistry and dynamics of chemotactic LTB synthesis.

Human leukotriene (LT) A hydrolase/aminopeptidase (LTAH) is a bifunctional enzyme that converts the highly unstable epoxide intermediate LTA into LTB, a potent leukocyte activating agent, while the aminopeptidase activity cleaves and inactivates the chemotactic tripeptide Pro-Gly-Pro. Here, we describe high-resolution crystal structures of LTAH complexed with LTA, providing the structural underpinnings of the enzyme's unique epoxide hydrolase (EH) activity, involving Zn, Y383, E271, D375, and two catalytic waters. The structures reveal that a single catalytic water is involved in both catalytic activities of LTAH, alternating between epoxide ring opening and peptide bond hydrolysis, assisted by E271 and E296, respectively.

Binding of Pro-Gly-Pro at the active site of leukotriene A4 hydrolase/aminopeptidase and development of an epoxide hydrolase selective inhibitor.

Leukotriene (LT) A4 hydrolase/aminopeptidase (LTA4H) is a bifunctional zinc metalloenzyme that catalyzes the committed step in the formation of the proinflammatory mediator LTB4. Recently, the chemotactic tripeptide Pro-Gly-Pro was identified as an endogenous aminopeptidase substrate for LTA4 hydrolase. Here, we determined the crystal structure of LTA4 hydrolase in complex with a Pro-Gly-Pro analog at 1.

Crystallographic analysis of 1,2,3-trichloropropane biodegradation by the haloalkane dehalogenase DhaA31.

Haloalkane dehalogenases catalyze the hydrolytic cleavage of carbon-halogen bonds, which is a key step in the aerobic mineralization of many environmental pollutants. One important pollutant is the toxic and anthropogenic compound 1,2,3-trichloropropane (TCP). Rational design was combined with saturation mutagenesis to obtain the haloalkane dehalogenase variant DhaA31, which displays an increased catalytic activity towards TCP.

Product formation controlled by substrate dynamics in leukotriene A4 hydrolase.

Leukotriene A4 hydrolase/aminopeptidase (LTA4H) (EC 3.3.2.

Crystallization and crystallographic analysis of the Rhodococcus rhodochrous NCIMB 13064 DhaA mutant DhaA31 and its complex with 1,2,3-trichloropropane.

Haloalkane dehalogenases hydrolyze carbon-halogen bonds in a wide range of halogenated aliphatic compounds. The potential use of haloalkane dehalogenases in bioremediation applications has stimulated intensive investigation of these enzymes and their engineering. The mutant DhaA31 was constructed to degrade the anthropogenic compound 1,2,3-trichloropropane (TCP) using a new strategy.

Crystallization and preliminary X-ray diffraction analysis of the wild-type haloalkane dehalogenase DhaA and its variant DhaA13 complexed with different ligands.

Haloalkane dehalogenases make up an important class of hydrolytic enzymes which catalyse the cleavage of carbon-halogen bonds in halogenated aliphatic compounds. There is growing interest in these enzymes owing to their potential use in environmental and industrial applications. The haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 can slowly detoxify the industrial pollutant 1,2,3-trichloropropane (TCP).

Atomic resolution studies of haloalkane dehalogenases DhaA04, DhaA14 and DhaA15 with engineered access tunnels.

The haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 is a bacterial enzyme that shows catalytic activity for the hydrolytic degradation of the highly toxic industrial pollutant 1,2,3-trichloropropane (TCP). Mutagenesis focused on the access tunnels of DhaA produced protein variants with significantly improved activity towards TCP. Three mutants of DhaA named DhaA04 (C176Y), DhaA14 (I135F) and DhaA15 (C176Y + I135F) were constructed in order to study the functional relevance of the tunnels connecting the buried active site of the protein with the surrounding solvent.

Pathways and mechanisms for product release in the engineered haloalkane dehalogenases explored using classical and random acceleration molecular dynamics simulations.

Eight mutants of the DhaA haloalkane dehalogenase carrying mutations at the residues lining two tunnels, previously observed by protein X-ray crystallography, were constructed and biochemically characterized. The mutants showed distinct catalytic efficiencies with the halogenated substrate 1,2,3-trichloropropane. Release pathways for the two dehalogenation products, 2,3-dichloropropane-1-ol and the chloride ion, and exchange pathways for water molecules, were studied using classical and random acceleration molecular dynamics simulations.

Crystals of DhaA mutants from Rhodococcus rhodochrous NCIMB 13064 diffracted to ultrahigh resolution: crystallization and preliminary diffraction analysis.

The enzyme DhaA from Rhodococcus rhodochrous NCIMB 13064 belongs to the haloalkane dehalogenases, which catalyze the hydrolysis of haloalkanes to the corresponding alcohols. The haloalkane dehalogenase DhaA and its variants can be used to detoxify the industrial pollutant 1,2,3-trichloropropane (TCP). Three mutants named DhaA04, DhaA14 and DhaA15 were constructed in order to study the importance of tunnels connecting the buried active site with the surrounding solvent to the enzymatic activity.