Structural analysis reveals a "molecular calipers" mechanism for a LATERAL ORGAN BOUNDARIES DOMAIN transcription factor protein from wheat.

LATERAL ORGAN BOUNDARIES DOMAIN (LBD) proteins, a family of plant-specific transcription factors harboring a conserved Lateral Organ Boundaries (LOB) domain, are regulators of plant organ development. Recent studies have unraveled additional pivotal roles of the LBD protein family beyond defining lateral organ boundaries, such as pollen development and nitrogen metabolism. The structural basis for the molecular network of LBD-dependent processes remains to be deciphered. Here, we solved the first structure of the homodimeric LOB domain of Ramosa2 from wheat (TtRa2LD) to 1.9 Å resolution. Our crystal structure reveals structural features shared with other zinc-finger transcriptional factors, as well as some features unique to LBD proteins. Formation of the TtRa2LD homodimer relied on hydrophobic interactions of its coiled-coil motifs. Several specific motifs/domains of the LBD protein were also involved in maintaining its overall conformation. The intricate assembly within and between the monomers determined the precise spatial configuration of the two zinc fingers that recognize palindromic DNA sequences. Biochemical, molecular modeling, and small-angle X-ray scattering experiments indicated that dimerization is important for cooperative DNA binding and discrimination of palindromic DNA through a molecular calipers mechanism. Along with previously published data, this study enables us to establish an atomic-scale mechanistic model for LBD proteins as transcriptional regulators in plants.

A 3'-5' exonuclease activity embedded in the helicase core domain of Candida albicans Pif1 helicase.

3'-5' exonucleases are frequently found to be associated to polymerases or helicases domains in the same enzyme or could function as autonomous entities. Here we uncovered that Candida albicans Pif1 (CaPif1) displays a 3'-5' exonuclease activity besides its main helicase activity. These two latter activities appear to reside on the same polypeptide and the new exonuclease activity could be mapped to the helicase core domain. We clearly show that CaPif1 displays exclusively exonuclease activity and unambiguously establish the directionality of the exonuclease activity as the 3'-to-5' polarity. The enzyme appears to follow the two-metal-ion driven hydrolyzing activity exhibited by most of the nucleases, as shown by its dependence of magnesium and also by the identification of aspartic residues. Interestingly, an excellent correlation could be found between the presence of the conserved residues and the exonuclease activity when testing activities on Pif1 enzymes from eight fungal organisms. In contrast to others proteins endowed with the double helicase/exonuclease functionality, CaPif1 differs in the fact that the two activities are embedded in the same helicase domain and not located on separated domains. Our findings may suggest a biochemical basis for mechanistic studies of Pif1 family helicases.

The cytochrome P450 2D6*10 genetic polymorphism alters postoperative analgesia.

The present study was aimed to investigate the effects of the cytochrome P450 (CYP) 2D6*10 genetic polymorphism on postoperative patient-controlled morphine usage. A total of 114 patients were selected, and 102 patients completed the study. Polymerase chain reaction-based restriction fragment length polymorphism (PCR-RFLP) was used to determine the CYP2D6*10 genotype, and patients were categorized into three groups according to CYP2D6 genotype: heterozygous (m/w), wild-type homozygous (w/w), and mutant homozygous (m/m). Total morphine usage and visual analogue score (VAS) were determined 72 hours after the operation and compared across the three genotype groups. Statistical methods used to analyze results were the χ(2) test, analysis of variance, and multiple linear regression analysis; P<0.05 was considered to be statistically significant. The cumulative use of morphine in the m/w group was significantly higher than that in the m/m group between T0.5 and T4h (P<0.05). There were no significant differences in the loading dose of morphine or VAS among the different genotypes within 72 hours of operation. Patients carrying the CYP2D6*10 m/w genotype required higher doses of morphine at T0.5~T4h compared to the m/m group, and therefore received a higher cumulative dose of morphine post-operation. This phenomenon may be due to a decreased ability to synthesize endogenous opioid peptide.

Selected polymorphisms of GSTP1 and TERT were associated with glioma risk in Han Chinese.

BACKGROUND: Current evidence suggests that a majority of the inherited risks play a major role in glioma susceptibility, and glioma is due to the co-inheritance of multiple low-risk variants. These variants can be identified through association studies including such as genome-wide association studies (GWAS), which has led the glioma epidemiology researchers to focus on identifying potential disease-causing factors. METHODS: We evaluated and validated 10 tag single nucleotide polymorphisms (tSNPs) in seven genes associated with glioma susceptibility in a Han Chinese population, including 301 glioma cases and 302 controls, using a multiplexed single nucleotide polymorphism (SNP) MassEXTEND assay. We ascertained the genotypic frequencies for each tSNP in control subjects were within Hardy-Weinberg equilibrium (HWE) using an exact test, and then compared the genotype and allele frequencies of glioma patients and control subjects using the χ2 test. We then applied three genetic models (dominant, recessive, and additive) using PLINK software to assess the association of each tSNP with glioma risk. RESULTS: We identified two tSNPs to be associated with glioma susceptibility (rs1695, GSTP1, P = 0.019; rs2853676, TERT, P = 0.039), which we confirmed using dominant and additive model analyses. The genotype “GA” for rs1695 was recognized to be a protective genotype for glioma (OR, 0.67; 95% CI, 0.47-0.96; P = 0.027), while the genotype “AG” for rs2853676 was shown to be a risk genotype for glioma (OR, 1.50; 95% CI, 1.05-2.15; P = 0.025). CONCLUSION: Our results, and those from previous studies, suggest potential genetic contributes for GSTP1 and TERT in glioma development.

[Clinical study on the antioxidant effect of methylene blue in vivo].

OBJECTIVE: To investigate the relationship of oxygen free radical (OFR) with per-oxidative injury of erythrocyte induced by intravenous procaine in vivo and the effect of methylene blue (MB) in removal of nitric oxide (NO) and peroxynitrite (ONOO(-)). METHODS: Forty patients undergoing elective surgery were divided randomly into intravenous procaine anesthesia (IPA) group and fentanyl group. Blood sample was taken before anesthesia (T0), 120 minutes (T1) and 180 minutes (T2) after IPA and 30 minutes after treatment with MB (1-2 mg/kg, T3) to determine the changes in the levels of NO, OFR, lipid peroxide (LPO), superoxide dismutase (SOD), catalase (CAT), NADH-Cyt b5-reductase (Cyt b5-R) and methemoglobin (MHb). RESULTS: Compared with T0, the levels of NO, OFR, LPO, MHb in IPA group were significantly increased at T1,T2. At same time SOD, CAT and Cyt b5-R were significantly decreased. NO, OFR, MHb, SOD, CAT and Cyt b5-R were all reduced to the normal levels at T3. No changes in any determined parameters in fentanyl group during anesthesia. CONCLUSION: It is indicated that the metabolites of procaine consist of a large quantity of NO:ONOO(-), producing per-oxidative injury to erythrocyte. MB is effective in eliminating OFR in vivo, protecting tissue cells. It may act as an antioxidant drug in the treatment of critical illness.