Screw head plugs increase the fatigue strength of stainless steel, but not of titanium, locking plates.

OBJECTIVES: Screw plugs have been reported to increase the fatigue strength of stainless steel locking plates. The objective of this study was to examine and compare this effect between stainless steel and titanium locking plates. METHODS: Custom-designed locking plates with identical structures were fabricated from stainless steel and a titanium alloy. Three types of plates were compared: type I unplugged plates; type II plugged plates with a 4 Nm torque; and type III plugged plates with a 12 Nm torque. The stiffness, yield strength, and fatigue strength of the plates were investigated through a four-point bending test. Failure analyses were performed subsequently. RESULTS: For stainless steel, type II and type III plates had significantly higher fatigue strength than type I plates. For titanium, there were no significant differences between the fatigue strengths of the three types of plates. Failure analyses showed local plastic deformations at the threads of screw plugs in type II and type III stainless steel plates but not in titanium plates. CONCLUSION: The screw plugs could increase the fatigue strength of stainless steel plates but not of titanium plates. Therefore, leaving screw holes open around fracture sites is recommended in titanium plates.Cite this article: L-W. Hung, C-K. Chao, J-R. Huang, J. Lin. Screw head plugs increase the fatigue strength of stainless steel, but not of titanium, locking plates. Bone Joint Res 2018;7:629-635. DOI: 10.1302/2046-3758.712.BJR-2018-0083.R1.

Restoration of intestinal function in an MPTP model of Parkinson's Disease.

Patients with Parkinson's disease often experience non-motor symptoms including constipation, which manifest prior to the onset of debilitating motor signs. Understanding the causes of these non-motor deficits and developing disease modifying therapeutic strategies has the potential to prevent disease progression. Specific neuronal subpopulations were reduced within the myenteric plexus of mice 21 days after intoxication by the intraperitoneal administration of MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) and was associated with a reduction in stool frequency, indicative of intestinal dysfunction. Oral administration of the divalent copper complex, Cu(II)(atsm), which has been shown to be neuroprotective and restore motor performance to MPTP lesioned mice, improved stool frequency and was correlated with restoration of neuronal subpopulations in the myenteric plexus of MPTP lesioned mice. Restoration of intestinal function was associated with reduced enteric glial cell reactivity and reduction of markers of inflammation. Therapeutics that have been shown to be neuroprotective in the central nervous system, such as Cu(II)(atsm), therefore also provide symptom relief and are disease modifying in the intestinal tract, suggesting that there is a common cause of Parkinson's disease pathogenesis in the enteric nervous system and central nervous system.

Discovery and spectroscopy of the young jovian planet 51 Eri b with the Gemini Planet Imager.

Directly detecting thermal emission from young extrasolar planets allows measurement of their atmospheric compositions and luminosities, which are influenced by their formation mechanisms. Using the Gemini Planet Imager, we discovered a planet orbiting the ~20-million-year-old star 51 Eridani at a projected separation of 13 astronomical units. Near-infrared observations show a spectrum with strong methane and water-vapor absorption. Modeling of the spectra and photometry yields a luminosity (normalized by the luminosity of the Sun) of 1.6 to 4.0 × 10(-6) and an effective temperature of 600 to 750 kelvin. For this age and luminosity, "hot-start" formation models indicate a mass twice that of Jupiter. This planet also has a sufficiently low luminosity to be consistent with the "cold-start" core-accretion process that may have formed Jupiter.

Pathway to the piezoelectronic transduction logic device.

The piezoelectronic transistor (PET) has been proposed as a transduction device not subject to the voltage limits of field-effect transistors. The PET transduces voltage to stress, activating a facile insulator-metal transition, thereby achieving multigigahertz switching speeds, as predicted by modeling, at lower power than the comparable generation field effect transistor (FET). Here, the fabrication and measurement of the first physical PET devices are reported, showing both on/off switching and cycling. The results demonstrate the realization of a stress-based transduction principle, representing the early steps on a developmental pathway to PET technology with potential to contribute to the IT industry.

Crystallization and preliminary X-ray crystallographic analysis of a Mycobacterium tuberculosis ferritin homolog, BfrB.

Mycobacterium tuberculosis (Mtb) is the causative agent of the deadly disease tuberculosis. Iron acquisition, regulation and storage are critical for the survival of this pathogen within a host. Thus, understanding the mechanisms of iron metabolism in Mtb will shed light on its pathogenic nature, as iron is important for infection. Ferritins are a superfamily of protein nanocages that function in both iron detoxification and storage, and Mtb contains both a predicted ferritin and a bacterioferritin. Here, the cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of the ferritin homolog (Mtb BfrB, Rv3841) is reported. An Mtb BfrB crystal grown at pH 6.5 using the hanging-drop vapor-diffusion technique diffracted to 2.50 Šresolution and belonged to space group C2, with unit-cell parameters a=226.2, b=226.8, c=113.7 Å, ß=94.7° and with 24 subunits per asymmetric unit. Furthermore, modeling the crystal structure of a homologous ferritin into a low-resolution small-angle X-ray scattering (SAXS) electron-density envelope is consistent with the presence of 24 subunits in the BfrB protein cage quaternary structure.

High throughput crystallography of TB drug targets.

Tuberculosis (TB) infects one-third of the world population. Despite 50 years of available drug treatments, TB continues to increase at a significant rate. The failure to control TB stems in part from the expense of delivering treatment to infected individuals and from complex treatment regimens. Incomplete treatment has fueled the emergence of multi-drug resistant (MDR) strains of Mycobacterium tuberculosis (Mtb). Reducing non-compliance by reducing the duration of chemotherapy will have a great impact on TB control. The development of new drugs that either kill persisting organisms, inhibit bacilli from entering the persistent phase, or convert the persistent bacilli into actively growing cells susceptible to our current drugs will have a positive effect. We are taking a multidisciplinary approach that will identify and characterize new drug targets that are essential for persistent Mtb. Targets are exposed to a battery of analyses including microarray experiments, bioinformatics, and genetic techniques to prioritize potential drug targets from Mtb for structural analysis. Our core structural genomics pipeline works with the individual laboratories to produce diffraction quality crystals of targeted proteins, and structural analysis will be completed by the individual laboratories. We also have capabilities for functional analysis and the virtual ligand screening to identify novel inhibitors for target validation. Our overarching goals are to increase the knowledge of Mtb pathogenesis using the TB research community to drive structural genomics, particularly related to persistence, develop a central repository for TB research reagents, and discover chemical inhibitors of drug targets for future development of lead compounds.

The TB structural genomics consortium: a resource for Mycobacterium tuberculosis biology.

The TB Structural Genomics Consortium is an organization devoted to encouraging, coordinating, and facilitating the determination and analysis of structures of proteins from Mycobacterium tuberculosis. The Consortium members hope to work together with other M. tuberculosis researchers to identify M. tuberculosis proteins for which structural information could provide important biological information, to analyze and interpret structures of M. tuberculosis proteins, and to work collaboratively to test ideas about M. tuberculosis protein function that are suggested by structure or related to structural information. This review describes the TB Structural Genomics Consortium and some of the proteins for which the Consortium is in the progress of determining three-dimensional structures.

Crystal structure of archaeal RNase HII: a homologue of human major RNase H.

BACKGROUND: RNases H are present in all organisms and cleave RNAs in RNA/DNA hybrids. There are two major types of RNases H that have little similarity in sequence, size and specificity. The structure of RNase HI, the smaller enzyme and most abundant in bacteria, has been extensively studied. However, no structural information is available for the larger RNase H, which is most abundant in eukaryotes and archaea. Mammalian RNase H participates in DNA replication, removal of the Okazaki fragments and possibly DNA repair. RESULTS: The crystal structure of RNase HII from the hypothermophile Methanococcus jannaschii, which is homologous to mammalian RNase H, was solved using a multiwavelength anomalous dispersion (MAD) phasing method at 2 A resolution. The structure contains two compact domains. Despite the absence of sequence similarity, the large N-terminal domain shares a similar fold with the RNase HI of bacteria. The active site of RNase HII contains three aspartates: Asp7, Asp112 and Asp149. The nucleotide-binding site is located in the cleft between the N-terminal and C-terminal domains. CONCLUSIONS: Despite a lack of any detectable similarity in primary structure, RNase HII shares a similar structural domain with RNase HI, suggesting that the two classes of RNases H have a common catalytic mechanism and possibly a common evolutionary origin. The involvement of the unique C-terminal domain in substrate recognition explains the different reaction specificity observed between the two classes of RNase H.

The crystal structure of the Rev binding element of HIV-1 reveals novel base pairing and conformational variability.

The crystal and molecular structure of an RNA duplex corresponding to the high affinity Rev protein binding element (RBE) has been determined at 2.1-A resolution. Four unique duplexes are present in the crystal, comprising two structural variants. In each duplex, the RNA double helix consists of an annealed 12-mer and 14-mer that form an asymmetric internal loop consisting of G-G and G-A noncanonical base pairs and a flipped-out uridine. The 12-mer strand has an A-form conformation, whereas the 14-mer strand is distorted to accommodate the bulges and noncanonical base pairing. In contrast to the NMR model of the unbound RBE, an asymmetric G-G pair with N2-N7 and N1-O6 hydrogen bonding, is formed in each helix. The G-A base pairing agrees with the NMR structure in one structural variant, but forms a novel water-mediated pair in the other. A backbone flip and reorientation of the G-G base pair is required to assume the RBE conformation present in the NMR model of the complex between the RBE and the Rev peptide.

Expression, crystallization and preliminary X-ray studies of the PDZ domain of Dishevelled protein.

Dishevelled (Dsh) protein is an important component of the Wnt signal-transduction pathway. It has three relatively conserved domains: DIX, PDZ and DEP. The PDZ domain of the Xenopus laevis homolog of Dsh, which consists of residues 254-348, was overexpressed as a soluble protein in Escherichia coli, purified and crystallized. The crystals were obtained by the vapor-diffusion method, using 1.4 M sodium formate as a precipitant. The crystals diffracted to 2.3 A resolution. The space group was determined to be P6(1)22 or P6(5)22, with unit-cell dimensions a = b = 95.9, c = 93.9 A.

Structural differences in D and L-monellin in the crystals of racemic mixture.

The racemic mixture of synthetic d and l-monellin has been crystallized, and its structure has been determined by X-ray crystallography at 1.9 A resolution. The crystal structure consists of two d and two l-monellin molecules in the P1 unit cell with a pseudo-centrosymmetrical arrangement. The final structure reveals small but significant structural differences between d and l-monellin in the same crystal. Possible reasons for these differences and their implications are discussed.

Structure-based assignment of the biochemical function of a hypothetical protein: a test case of structural genomics.

Many small bacterial, archaebacterial, and eukaryotic genomes have been sequenced, and the larger eukaryotic genomes are predicted to be completely sequenced within the next decade. In all genomes sequenced to date, a large portion of these organisms' predicted protein coding regions encode polypeptides of unknown biochemical, biophysical, and/or cellular functions. Three-dimensional structures of these proteins may suggest biochemical or biophysical functions. Here we report the crystal structure of one such protein, MJ0577, from a hyperthermophile, Methanococcus jannaschii, at 1.7-A resolution. The structure contains a bound ATP, suggesting MJ0577 is an ATPase or an ATP-mediated molecular switch, which we confirm by biochemical experiments. Furthermore, the structure reveals different ATP binding motifs that are shared among many homologous hypothetical proteins in this family. This result indicates that structure-based assignment of molecular function is a viable approach for the large-scale biochemical assignment of proteins and for discovering new motifs, a basic premise of structural genomics.

Structure of an enantiomeric protein, D-monellin at 1.8 A resolution.

The D-enantiomer of a potently sweet protein, monellin, has been crystallized and analyzed by X-ray crystallography at 1.8 A resolut ion. Two crystal forms (I and II) appeared under crystallization conditions similar, but not identical, to the crystallization conditions of natural L-monellin. There are four molecules per asymmetric unit in crystal form I and one in crystal form II. Crystal form I is not reproducible and is equivalent to that of monoclinic L-monellin. Intermonomer contacts in crystal form II are very different from those found in natural L-monellin crystals. The backbone trace of D-monellin resembles very closely the mirror image of that of L-monellin, but the N- and C-terminus backbones as well as several side-chain conformations of D-monellin are different from those of natural L-monellin. Most of these apparent differences may be attributable to the crystal packing differences.

Purification, crystallization and preliminary X-ray crystallographic analysis of Pyrococcus furiosus DNA polymerase.

DNA polymerase gene from the hyperthermophilic Archaeon Pyrococcus furiosus has been cloned and the protein overexpressed in Escherichia coli to produce an active enzyme. The purified protein was crystallized from 0.08 M ammonium sulfate, 0.05 M Na-cacodylate, pH 6.5, 0.15%(v/v) NP40, 0.05%(v/v) Tween 20 and 4.5%(w/v) polyethylene glycol 6000 by the vapour-diffusion method. The orthorhombic crystals had unit-cell dimensions of a = 92.5, b = 125.4, c = 192.1 A; alpha = beta = gamma = 90 degrees. The crystals diffracted beyond 4 A on a 1.08 A synchrotron radiation source.

Crystal structures of eukaryotic translation initiation factor 5A from Methanococcus jannaschii at 1.8 A resolution.

Eukaryotic translation initiation factor 5A (eIF-5A) is a ubiquitous protein found in all eukaryotic cells. The protein is closely associated with cell proliferation in the G1-S stage of the cell cycle. Recent findings show that the eIF-5A proteins are highly expressed in tumor cells and act as a cofactor of the Rev protein in HIV-1-infected cells. The mature eIF is the only protein known to have the unusual amino acid hypusine, a post-translationally modified lysine. The crystal structure of eIF-5A from Methanococcus jannaschii (MJ eIF-5A) has been determined at 1.9 A and 1.8 A resolution in two crystal forms by using the multiple isomorphous replacement method and the multiwavelength anomalous diffraction method for the first crystal form and the molecular replacement method for the second crystal form. The structure consists of two folding domains, one of which is similar to the oligonucleotide-binding domain found in the prokaryotic cold shock protein and the translation initiation factor IF1 despite the absence of any significant sequence similarities. The 12 highly conserved amino acid residues found among eIF-5As include the hypusine site and form a long protruding loop at one end of the elongated molecule.

Structure of an RNA internal loop consisting of tandem C-A+ base pairs.

The crystal structure of the RNA octamer 5'-CGC(CA)GCG-3' has been determined from X-ray diffraction data to 2.3 A resolution. In the crystal, this oligomer forms a self-complementary double helix in the asymmetric unit. Tandem non-Watson-Crick C-A and A-C base pairs comprise an internal loop in the middle of the duplex, which is incorporated with little distortion of the A-form double helix. From the geometry of the C-A base pairs, it is inferred that the adenosine imino group is protonated and donates a hydrogen bond to the carbonyl group of the cytosine. The wobble geometry of the C-A+ base pairs is very similar to that of the common U-G non-Watson-Crick pair.

Electron transfer by domain movement in cytochrome bc1.

The cytochrome bc1 is one of the three major respiratory enzyme complexes residing in the inner mitochondrial membrane. Cytochrome bc1 transfers electrons from ubiquinol to cytochrome c and uses the energy thus released to form an electrochemical gradient across the inner membrane. Our X-ray crystal structures of the complex from chicken, cow and rabbit in both the presence and absence of inhibitors of quinone oxidation, reveal two different locations for the extrinsic domain of one component of the enzyme, an iron-sulphur protein. One location is close enough to the supposed quinol oxidation site to allow reduction of the Fe-S protein by ubiquinol. The other site is close enough to cytochrome c1 to allow oxidation of the Fe-S protein by the cytochrome. As neither location will allow both reactions to proceed at a suitable rate, the reaction mechanism must involve movement of the extrinsic domain of the Fe-S component in order to shuttle electrons from ubiquinol to cytochrome c1. Such a mechanism has not previously been observed in redox protein complexes.

Crystal structure of the ATP-binding subunit of an ABC transporter.

ABC transporters (also known as traffic ATPases) form a large family of proteins responsible for the translocation of a variety of compounds across membranes of both prokaryotes and eukaryotes. The recently completed Escherichia coli genome sequence revealed that the largest family of paralogous E. coli proteins is composed of ABC transporters. Many eukaryotic proteins of medical significance belong to this family, such as the cystic fibrosis transmembrane conductance regulator (CFTR), the P-glycoprotein (or multidrug-resistance protein) and the heterodimeric transporter associated with antigen processing (Tap1-Tap2). Here we report the crystal structure at 1.5 A resolution of HisP, the ATP-binding subunit of the histidine permease, which is an ABC transporter from Salmonella typhimurium. We correlate the details of this structure with the biochemical, genetic and biophysical properties of the wild-type and several mutant HisP proteins. The structure provides a basis for understanding properties of ABC transporters and of defective CFTR proteins.

Crystallization and preliminary X-ray analysis of D-monellin.

D-Monellin is a chemically synthesized protein composed of all D-amino acids. It has an amino-acid sequence identical to L-monellin, a natural protein with potent sweetness. Two crystal forms of D-monellin were obtained. Both crystals were grown under conditions similiar to those used to crystallize natural L-monellin. Crystal form I has similar, but not identical, cell parameters to natural L-monellin and diffracts to 2.7 A resolution. Crystal form II is very different and diffracts to 1.7 A resolution using synchrotron radiation.