Precipitant-Free Lysozyme Crystals Grown by Centrifugal Concentration Reveal Structural Changes

The three-dimensional (3D) structure of a protein molecule in its crystal need not correspond to that found in vivo in many cases, since we usually crystallize protein molecules using precipitants (salts, organic solvents, polymeric electrolytes, etc.), and the precipitants are often incorporated into crystals along with the protein molecules. Although precipitant-free crystallization methods would solve these problems, such methods had not yet been established. We have achieved a novel precipitant-free crystallization method by liquid-liquid phase separation during the centrifugal concentration of lysozyme in ultra-pure water. In the 3D structure of the precipitant-free crystal, lysozyme loses a sodium cation and changes the position of Ser 72. Deionization of the solution also appears to induce a change in the position of Asp 101 and an increase in the activity of lysozyme

Giant High-Flow Type Pulmonary Arteriovenous Malformation: Coil Embolization with Flow Control by Balloon Occlusion and an Anchored Detachable Coil

Pulmonary arteriovenous malformations (PAVMs) are often treated by pushable fibered or non-fibered microcoils, using an anchor or scaffold technique or with an Amplatzer plug through a guiding sheath. When performing percutaneous transcatheter microcoil embolization, there is a risk of coil migration, particularly with high-flow type PAVMs. The authors report on a unique treatment in a patient with a giant high-flow PAVM whose nidus had a maximum diameter of 6 cm. A detachable coil, not detached from a delivery wire (an anchored detachable coil), was first placed in the feeding artery under flow control by balloon occlusion, and then multiple microcoils were packed proximally to the anchored detachable coil. After confirming the stability of the microcoils during a gradual deflation of the balloon, we finally released the first detachable coil. The nidus was reduced in size to 15 mm at one year postoperatively

Identification of effector candidate genes of Rhizoctonia solani AG-1 IA expressed during infection in Brachypodium distachyon

Rhizoctonia solani is a necrotrophic phytopathogen belonging to basidiomycetes. It causes rice sheath blight which inflicts serious damage in rice production. The infection strategy of this pathogen remains unclear. We previously demonstrated that salicylic acid-induced immunity could block R. solani AG-1 IA infection in both rice and Brachypodium distachyon. R. solani may undergo biotrophic process using effector proteins to suppress host immunity before necrotrophic stage. To identify pathogen genes expressed at the early infection process, here we developed an inoculation method using B. distachyon which enables to sample an increased amount of semi-synchronous infection hyphae. Sixty-one R. solani secretory effector-like protein genes (RsSEPGs) were identified using in silico approach with the publicly available gene annotation of R. solani AG-1 IA genome and our RNA-sequencing results obtained from hyphae grown on agar medium. Expression of RsSEPGs was analyzed at 6, 10, 16, 24, and 32 h after inoculation by a quantitative reverse transcription-polymerase chain reaction and 52 genes could be detected at least on a single time point tested. Their expressions showed phase-specific patterns which were classified into 6 clusters. The 23 RsSEPGs in the cluster 1-3 and 29 RsSEPGs in the cluster 4-6 are expected to be involved in biotrophic and necrotrophic interactions, respectively

Population pharmacokinetics and pharmacodynamics of sitafloxacin in patients with community-acquired respiratory tract infections

An optimal dosage regimen of sitafloxacin was considered based on a pharmacokinetics and pharmacodynamics (PK–PD) analysis in patients with community-acquired respiratory tract infections (RTI). A population pharmacokinetic analysis of sitafloxacin was conducted using clinical data of five clinical pharmacology studies and one clinical PK–PD study in patients with RTIs. The pharmacokinetic parameters in individual patients were estimated by the Bayesian method to examine any correlation between pharmacokinetics and bacteriological efficacy. Efficacy data were obtained from the clinical PK–PD study, in which 50 or 100 mg sitafloxacin was administered twice daily for 7 days. In addition, an efficacy was simulated for a hypothetical dose regimen of 100 mg once daily. The fAUC(0–24h)/MIC and the fC(max)/MIC of sitafloxacin at a dose of 50 mg twice daily were 117.5 ± 78.0 and 7.3 ± 4.7 (mean ± SD), respectively. As a result of the univariate logistic regression analysis, the larger the value of fAUC(0–24h)/MIC or fC(max)/MIC becomes, the higher the bacteriological efficacies. The eradication rates for fAUC(0–24h)/MIC ≥ 30 and for fC(max)/MIC ≥ 2 were 96.4 % and 96.3 %, respectively. The PK–PD target values of sitafloxacin for the treatment of mild to moderate RTIs were considered to be fAUC(0–24h)/MIC ≥ 30 and fC(max)/MIC ≥ 2. The PK–PD parameters at the regimen of 50 or 100 mg twice daily in patients with RTIs reached the target values. Furthermore, a 100 mg once-daily regimen was expected to show similar efficacy based on the PK–PD simulations

A Polymorphism in MAPKAPK3 Affects Response to Interferon Therapy for Chronic Hepatitis C

Background & Aims: This study aimed to identify host single nucleotide polymorphisms (SNPs) that are associated with the efficacy of interferon (IFN) therapy in patients with chronic hepatitis C. Methods: We examined whether 116 tagging-SNPs from 13 genes that are involved in type I IFN signaling associate with the outcome of IFN therapy in Japanese case-control groups; the study included 468 sustained responders and 587 nonresponders. Results: We identified 2 SNPs (rs3792323 [A/T] and rs616589 [G/A]), located in intron 2 of mitogen-activated protein kinase-activated protein kinase 3 (MAPKAIPK3) that were associated with the outcome of IFN therapy in patients infected with hepatitis C virus (HCV) genotype 1b (P = 4.6 X 10(-5) and 4.8 X 10(-5), respectively). The 2 SNPs were in strong linkage disequilibrium and multivariate logistic regression analysis showed that rs3792323 is an independent factor associated with the IFN efficacy (genotype 1b; P =.0011). MAPKAPK3 is a kinase involved in the mitogen and stress responses, but the biological significance of MAPKAPK3 in IFN responses is poorly understood. By using an allele-specific transcript quantification assay in liver biopsy, we showed that allelespecific expression of MAPKAPK3 messenger RNA, corresponding to the risk allele for nonresponse, was significantly higher than that of the other allele. Luciferase reporter assay data indicated that overexpression of MAPKAPK3 inhibits IFN-alfa-induced gene transcription via IFN-stimulated response element and IFN gamma-activated site. Conclusions: The SNP rs3792323 in MAPKAPK3 associates with the outcome of IFN therapy in patients with HCV genotype 1b. Our functional analyses indicate that MAPKAPK3 inhibits IFN-alfa-induced antiviral activity