Parasite-mediated predation determines infection in a complex predator-prey-parasite system.

The interplay of host-parasite and predator-prey interactions is critical in ecological dynamics because both predators and parasites can regulate communities. But what is the prevalence of infected prey and predators when a parasite is transmitted through trophic interactions considering stochastic demographic changes? Here, we modelled and analysed a complex predator-prey-parasite system, where parasites are transmitted from prey to predators. We varied parasite virulence and infection probabilities to investigate how those evolutionary factors determine species' coexistence and populations' composition. Our results show that parasite species go extinct when the infection probabilities of either host are small and that success in infecting the final host is more critical for the survival of the parasite. While our stochastic simulations are consistent with deterministic predictions, stochasticity plays an important role in the border regions between coexistence and extinction. As expected, the proportion of infected individuals increases with the infection probabilities. Interestingly, the relative abundances of infected and uninfected individuals can have opposite orders in the intermediate and final host populations. This counterintuitive observation shows that the interplay of direct and indirect parasite effects is a common driver of the prevalence of infection in a complex system.

The HD-Domain Metalloprotein Superfamily: An Apparent Common Protein Scaffold with Diverse Chemistries.

The histidine-aspartate (HD)-domain protein superfamily contains metalloproteins that share common structural features but catalyze vastly different reactions ranging from oxygenation to hydrolysis. This chemical diversion is afforded by (i) their ability to coordinate most biologically relevant transition metals in mono-, di-, and trinuclear configurations, (ii) sequence insertions or the addition of supernumerary ligands to their active sites, (iii) auxiliary substrate specificity residues vicinal to the catalytic site, (iv) additional protein domains that allosterically regulate their activities or have catalytic and sensory roles, and (v) their ability to work with protein partners. More than 500 structures of HD-domain proteins are available to date that lay out unique structural features which may be indicative of function. In this respect, we describe the three known classes of HD-domain proteins (hydrolases, oxygenases, and lyases) and identify their apparent traits with the aim to portray differences in the molecular details responsible for their functional divergence and reconcile existing notions that will help assign functions to yet-to-be characterized proteins. The present review collects data that exemplify how nature tinkers with the HD-domain scaffold to afford different chemistries and provides insight into the factors that can selectively modulate catalysis.

Neighbors help neighbors control urban mosquitoes.

The worldwide spread of invasive Aedes mosquitoes and arboviral disease, have renewed the pressure for effective and sustainable urban mosquito control. We report on the success of a model we are confident will usher in a new era of urban mosquito control. The key innovation is the mobilization of neighbors guided by scientific advisors, an approach we termed Citizen Action through Science (Citizen AcTS). This approach was tested in a NE US town of approximately 1,000 residential yards infested with the invasive Asian tiger mosquito, Aedes albopictus, a major nuisance arboviral vector. We report a highly significant reduction in biting pressure that was maintained over time, and establish the thresholds needed for success. The Citizen AcTS model rejects the top-down approach consistently associated with intervention failures. Instead, it works through respectful exchanges among scientists and residents that lead to trust and individual 'buy-in' and transferring program ownership to the community.

Structure Determination of Mycobacterium tuberculosis Serine Protease Hip1 (Rv2224c).

The Mycobacterium tuberculosis (Mtb) serine protease Hip1 (hydrolase important for pathogenesis; Rv2224c) promotes tuberculosis (TB) pathogenesis by impairing host immune responses through proteolysis of a protein substrate, Mtb GroEL2. The cell surface localization of Hip1 and its immunomodulatory functions make Hip1 a good drug target for new adjunctive immune therapies for TB. Here, we report the crystal structure of Hip1 to a resolution of 2.6 Å and the kinetic studies of the enzyme against model substrates and the protein GroEL2. The structure shows a two-domain protein, one of which contains the catalytic residues that are the signature of a serine protease. Surprisingly, a threonine is located within the active site close enough to hydrogen bond with the catalytic residues Asp463 and His490. Mutation of this residue, Thr466, to alanine established its importance for function. Our studies provide insights into the structure of a member of a novel family of proteases. Knowledge of the Hip1 structure will aid in designing inhibitors that could block Hip1 activity.

Evidence of the participation of remote residues in the catalytic activity of Co-type nitrile hydratase from Pseudomonas putida.

Active sites may be regarded as layers of residues, whereby the residues that interact directly with substrate also interact with residues in a second shell and these in turn interact with residues in a third shell. These residues in the second and third layers may have distinct roles in maintaining the essential chemical properties of the first-shell catalytic residues, particularly their spatial arrangement relative to the substrate binding pocket, and their electrostatic and dynamic properties. The extent to which these remote residues participate in catalysis and precisely how they affect first-shell residues remains unexplored. To improve our understanding of the roles of second- and third-shell residues in catalysis, we used THEMATICS to identify residues in the second and third shells of the Co-type nitrile hydratase from Pseudomonas putida (ppNHase) that may be important for catalysis. Five of these predicted residues, and three additional, conserved residues that were not predicted, have been conservatively mutated, and their effects have been studied both kinetically and structurally. The eight residues have no direct contact with the active site metal ion or bound substrate. These results demonstrate that three of the predicted second-shell residues (α-Asp164, ß-Glu56, and ß-His147) and one predicted third-shell residue (ß-His71) have significant effects on the catalytic efficiency of the enzyme. One of the predicted residues (α-Glu168) and the three residues not predicted (α-Arg170, α-Tyr171, and ß-Tyr215) do not have any significant effects on the catalytic efficiency of the enzyme.

Zinc coordination geometry and ligand binding affinity: the structural and kinetic analysis of the second-shell serine 228 residue and the methionine 180 residue of the aminopeptidase from Vibrio proteolyticus.

The chemical properties of zinc make it an ideal metal to study the role of coordination strain in enzymatic rate enhancement. The zinc ion and the protein residues that are bound directly to the zinc ion represent a functional charge/dipole complex, and polarization of this complex, which translates to coordination distortion, may tune electrophilicity, and hence, reactivity. Conserved protein residues outside of the charge/dipole complex, such as second-shell residues, may play a role in supporting the electronic strain produced as a consequence of functional polarization. To test the correlation between charge/dipole polarity and ligand binding affinity, structure-function studies were carried out on the dizinc aminopeptidase from Vibrio proteolyticus. Alanine substitutions of S228 and M180 resulted in catalytically diminished enzymes whose crystal structures show very little change in the positions of the metal ions and the protein residues. However, more detailed inspections of the crystal structures show small positional changes that account for differences in the zinc ion coordination geometry. Measurements of the binding affinity of leucine phosphonic acid, a transition state analogue, and leucine, a product, show a correlation between coordination geometry and ligand binding affinity. These results suggest that the coordination number and polarity may tune the electrophilicity of zinc. This may have provided the evolving enzyme with the ability to discriminate between reaction coordinate species.

Structural and thermodynamic studies of simple aldose reductase-inhibitor complexes.

The competitive inhibition constants of series of inhibitors related to phenylacetic acid against both wild-type and the doubly mutanted C298A/W219Y aldose reductase have been measured. Van't Hoff analysis shows that these acids bind with an enthalpy near -6.8 kcal/mol derived from the electrostatic interactions, while the 100-fold differences in binding affinity appear to be largely due to entropic factors that result from differences in conformational freedom in the unbound state. These temperature studies also point out the difference between substrate and inhibitor binding. X-ray crystallographic analysis of a few of these inhibitor complexes both confirms the importance of a previously described anion binding site and reveals the hydrophobic nature of the primary binding site and its general plasticity. Based on these results, N-glycylthiosuccinimides were synthesized to demonstrate their potential in studies that probe distal binding sites. Reduced alpha-lipoic acid, an anti-oxidant and therapeutic for diabetic complications, was shown to bind aldose reductase with a binding constant of 1 microM.

A novel dnaC mutation that suppresses priB rep mutant phenotypes in Escherichia coli K-12.

The loading of a replisome in prokaryotic and eukaryotic cells at an origin of DNA replication and during replication restart is a highly ordered and regulated process. During replication restart in Escherichia coli, the PriA, PriB, PriC, DnaT and Rep proteins form multiple pathways that bind to repaired replication forks. These complexes are then recognized by DnaC as sites to load DnaB, the replicative helicase. Several dnaC mutations have been isolated that suppress phenotypes of some replication restart mutants. A new dnaC mutation (dnaC824) is reported here that efficiently suppresses priB rep mutant phenotypes. Furthermore, it is shown that dnaC824 will suppress phenotypes of priB priA300, rep priA300 and priB priC strains. Unlike other dnaC suppressors, it can only weakly suppress the absence of priA. Others have reported a different type of dnaC mutation, dnaC1331, is able to mimic priB mutant phenotypes. This is supported herein by showing that like dnaC1331, a priB mutation is synthetically lethal with a dam mutation and this can be rescued by a mutH mutation. Furthermore, priB dam lethality can also be suppressed by dnaC824. Like a priB mutation, a dnaC1331 mutation causes a priA2::kan-like phenotype when combined with priA300. Lastly, we show that dnaC824 is dominant to wild type and that dnaC1331 is recessive to wild type. Several models are discussed for the action of these mutant dnaC proteins in replication restart.

The structure of Apo R268A human aldose reductase: hinges and latches that control the kinetic mechanism.

Aldose reductase (AR) catalyzes the NADPH-dependent reduction of glucose and other sugars to their respective sugar alcohols. The NADP+/NADPH exchange is the rate-limiting step for this enzyme and contributes in varying degrees to the catalytic rates of other aldo-keto reductase superfamily enzymes. The mutation of Arg268 to alanine in human recombinant AR removes one of the ligands of the C2-phosphate of NADP+ and markedly reduces the interaction of the apoenzyme with the nucleotide. The crystal structure of human R268A apo-aldose reductase determined to a resolution of 2.1 A is described. The R268A mutant enzyme has similar kinetic parameters to the wild-type enzyme for aldehyde substrates, yet has greatly reduced affinity for the nucleotide substrate which greatly facilitates its crystallization in the apoenzyme form. The apo-structure shows that a high temperature factor loop (between residues 214 and 226) is displaced by as much as 17 A in a rigid body fashion about Gly213 and Ser226 in the absence of the nucleotide cofactor as compared to the wild-type holoenzyme structure. Several factors act to stabilize the NADPH-holding loop in either the 'open' or 'closed' conformations: (1) the presence and interactions of the nucleotide cofactor, (2) the residues surrounding the Gly213 and Ser226 hinges which form unique hydrogen bonds in the 'open' or 'closed' structure, and (3) the Trp219 "latch" residue which interacts with an arginine residue, Arg293, in the 'open' conformation or with a cysteine residue, Cys298, in the 'closed' conformation. Several mutations in and around the high temperature factor loop are examined to elucidate the role of the loop in the mechanism by which aldose reductase binds and releases its nucleotide substrate.