Investigation of liver-targeted peripheral focused ultrasound stimulation (pFUS) and its effect on glucose homeostasis and insulin resistance in type 2 diabetes mellitus: a proof of concept, phase 1 trial.

BACKGROUND: Mechanical waves produced by ultrasound pulses have been shown to activate mechanosensitive ion channels and modulate peripheral nerves. However, while peripheral ultrasound neuromodulation has been demonstrated in vitro and in pre-clinical models, there have been few reports of clinical tests. AIM: We modified a diagnostic imaging system for ultrasound neuromodulation in human subjects. We report the first safety and feasibility outcomes in subjects with type 2 diabetes (T2D) mellitus and discuss these outcomes in relation to previous pre-clinical results. DESIGN: The study was performed as an open label feasibility study to assess the effects of hepatic ultrasound (targeted to the porta hepatis) on glucometabolic parameters in subjects with T2D. Stimulation (peripheral focused ultrasound stimulation treatment) was performed for 3 days (i.e. 15 min per day), preceded by a baseline examination and followed by a 2-week observation period. METHODS: Multiple metabolic assays were employed including measures of fasting glucose and insulin, insulin resistance and glucose metabolism. The safety and tolerability were also assessed by monitoring adverse events, changes in vital signs, electrocardiogram parameters and clinical laboratory measures. RESULTS AND CONCLUSION: We report post-pFUS trends in several outcomes that were consistent with previous pre-clinical findings. Fasting insulin was lowered, resulting in a reduction of HOMA-IR scores (P-value 0.01; corrected Wilcoxon signed-rank test). Additional safety and exploratory markers demonstrated no device-related adverse impact of pFUS. Our findings demonstrate that pFUS represents a promising new treatment modality that could be used as a non-pharmaceutical adjunct or even alternative to current drug treatments in diabetes.

Machine learning based de-noising of electron back scatter patterns of various crystallographic metallic materials fabricated using laser directed energy deposition.

A novel machine learning (ML) method of refining noisy Electron Back Scatter Patterns (EBSP) is proposed. For this, conditional generative adversarial networks (c-GAN) have been employed. The problem of de-noising the EBSPs was formulated as an image translation task conditioned on the input images to get refined/denoised output of EBSPs which can be indexed using conventional Hough transform based indexing algorithms. The ML model was trained using 10,000 EBSPs acquired under different settings for additively manufactured FCC, BCC and HCP alloy samples ensuring enough diversity and complexity in training data set. Pairs of noisy and corresponding optimal EBSPs were acquired by suitable tweaking of the EBSP acquisition parameters such as beam defocus, pattern binning and EBSD camera exposure duration. The trained model has brought out significant improvement in EBSD indexing success rate on test data, accompanied by betterment of indexing accuracy, quantified through 'pattern fit'. Complete automation of the EBSP refinement was demonstrated where in entire EBSD scan data can be fed to the model to get the refined EBSPs from which high quality EBSD data can be obtained.

Hyperhomocysteinemia-related serum metabolome alterations not normalized by short-term folic acid treatment.

INTRODUCTION: Hyperhomocysteinemia (HHCys) is an independent risk factor for various diseases such as cardiovascular diseases, Alzheimer's, and cancers. Folate deficiency is one of the significant reasons for HHCys. However, it is not known whether folate deficiency with HHCys is associated with any serum metabolites. OBJECTIVES: Our objective was to identify the metabolic alterations in people having folate deficiency with HHCys and check whether a short-term folic acid therapy could reverse those metabolic changes. METHODS: The study enrolled 34 participants aged between 18 and 40 years having folate deficiency (< 4.6 ng/mL) with HHCys (> 15 µmol/L) and 21 normal healthy individuals. A short-term intervention of oral folic acid (5 mg/day) was done in the HHCys group for 30 days. Untargeted metabolomics analysis of serum was performed in all study subjects before and after the folic acid treatment. Different univariate methods and the multivariable-adjusted linear regression models were employed to determine an association between homocysteine level and metabolite profile. RESULTS: Metabolomics analysis data showed that many metabolites involved in the biochemical pathways of lipid metabolisms such as polyunsaturated fatty acids, glycerolipids, and phospholipids were downregulated in the HHCys group. Short-term oral folic acid therapy significantly reduced their serum homocysteine level. However, the metabolic pathway alterations observed in folate-deficient HHCys-condition were unaltered even after the folic acid treatment. CONCLUSIONS: Our study revealed that people who have a folic acid deficiency with HHCys have an altered metabolite profile related to lipid metabolism, which cannot be reversed by short-term folic acid therapy.

Stereotactic body radiation therapy for early-stage primary lung cancer, is an active breath coordinator necessary? An audit from a tertiary cancer care center.

CONTEXT: The hypofractionated stereotactic body radiation therapy (SBRT) has emerged as a safe and effective treatment modality for early-stage nonsmall cell lung carcinoma. AIMS: An audit SBRT in primary lung cancer treated in our center with and without an active breath coordinator (ABC) was undertaken to evaluate its impact on target volumes and clinical outcomes. SETTINGS AND DESIGN: This was an observational study. MATERIALS AND METHODS: Nine patients with lung carcinoma were treated from January 2014 to August 2016. Five patients were simulated using ABC and four patients with free breathing. Volumetric modulated arc therapy plans were generated using Monaco treatment planning software. Three patients were treated with a dose of 54 Gy in three fractions and six patients with a dose of 48 Gy in four fractions. STATISTICAL ANALYSIS USED: The statistical analysis was performed using Kaplan-Meier survival. RESULTS: The mean planning target volumes (PTV) in ABC and free breathing groups were 42.19cc and 60.17cc, respectively. The mean volume of lung receiving 20, 10, and 5 Gy (V20, V10and V5) in ABC group were 5.37cc, 10.49cc, and 18.45cc whereas in free breathing 6.63cc, 12.74cc, and 20.64cc, respectively. At a median follow-up of 18 months, there were three local recurrences. No significant toxicity occurred in our series. CONCLUSION: Our initial results show that SBRT is well tolerated with good local control. Although the PTV volume and irradiated normal lung volume was higher in this group compared to ABC group, this did not translate to any added clinical toxicity.

Stereotactic body radiation therapy for hepatocellular carcinoma: Experience from a tertiary cancer care center in India.

AIM: The aim of this retrospective study is to assess the toxicity and tumor response of stereotactic body radiation therapy (SBRT) protocol for hepatocellular carcinoma (HCC) in our institution. BACKGROUND: Hepatocellular cancer is one of the leading cancers among men in India. In recent years, SBRT has emerged as a promising tool in the treatment of HCC. MATERIALS AND METHODS: Ten patients diagnosed as HCC with Barcelona Clinic Liver Cancer Stage B and C, treated with SBRT technique from January 2013 to December 2016, were included in this study. SBRT was delivered using 6 MV photons with volumetric modulated arc therapy. Acute and late toxicities were graded, and tumor response was assessed using response evaluation criteria in solid tumors criteria. Kaplan-Meier curves were generated for progression-free survival (PFS) and overall survival (OS). RESULTS: The median age was 61.5 (52-69) years. The radiation dose ranged from 35 Gy to 60 Gy. All patients obtained partial response during assessment at 3 months after completion of treatment. The median PFS is 8 months (95% confidence interval [CI] - 5.22-10.77 months). The median OS is 51 months (95% CI - 17.64-65.10 months). The OS at 1 and 2 years is 75% and 57%, respectively. CONCLUSIONS: SBRT is well tolerated by our patients. The 1- and 2-year OS of 75% and 57% is consistent with other prospective and retrospective SBRT studies from the literature.

Preoperative long-course chemoradiation for localized rectal cancer: A retrospective comparison of response and outcome between 5-fluorouracil/leucovorin versus capecitabine.

BACKGROUND: Preoperative concurrent chemoradiation therapy (CRT) with either capecitabine or 5-florouracil/leucovorin (5 FU/LV) is the standard of care in locally advanced rectal cancer (LARC). Literature comparing the toxicity and response of these two regimens in Indian patients is sparse. Our objective was to compare the pathological response (PR) and clinical outcome of capecitabine versus 5 FU/LV in CRT for LARC. MATERIALS AND METHODS: Sixty patients with LARC treated with preoperative CRT with capecitabine or 5FU/LV from January 2009 to May 2014 were analyzed. Ryan's tumor regression grading was used for PR assessment and tumor downstaging was defined as a reduction in the T and N stages by at least one level. Toxicity was assessed with RTOG acute toxicity assessment criteria and CTCAE 4.0 version. Statistical analysis was done using IBM SPSS 20 software. Percentage of patients with respect to response rates and toxicities was computed in each of the treatment groups. To test the statistical significance of the difference in PR rates and toxicities between the two groups, Chi-square test was used. Kaplan-Meier estimate of survival rate was computed for each group. To test the statistical significance of the difference in survival rate, the log-rank test was applied. RESULTS AND CONCLUSION: The two groups (5 FU/LV vs. capecitabine) were comparable with respect to pathological complete response (20% vs. 24%), pathological downstaging (76% vs. 69%), sphincter preservation rates, and acute complication rates. Both regimens were well tolerated. Overall survival and disease-free survival also did not show a statistically significant difference between the two groups (P values 0.720 and 0.255, respectively). In summary, our analysis showed the equivalence of both regimens in the preoperative CRT setting.

Pediatric brain tumors: An analysis of 5 years of data from a tertiary cancer care center, India.

BACKGROUND: Pediatric brain tumors are the most common solid tumors in children and a leading cause of mortality and morbidity in children worldwide. Even though there are enough data about the epidemiology of pediatric brain tumors in western population, there are only a few reports from developing countries like India. AIMS: To study the epidemiological patterns of brain tumors in children, to study the patterns of care, and to assess the treatment response. MATERIALS AND METHODS: A retrospective epidemiological approach is used. The records of children <18 years registered in our department from August 2006 to July 2011 diagnosed as primary brain tumors are selected. Data regarding age, sex, site of the tumor, clinical features, histology, treatment plan, and treatment response are collected. The World Health Organization classification of neoplasms was adopted. RESULTS: Of 250 cases, females (57%) slightly outnumbered males. The present study revealed that astrocytoma (52%) is the most common brain tumor in childhood. Surgery was the main modality of treatment. Chemotherapy was given to 16% of patients. Even though radiation therapy was offered to 74% of patients, only 42% completed radiotherapy. There was subjective clinical improvement in 68% of patient population after treatment. CONCLUSIONS: This is the second study from Tamil Nadu that deals with epidemiology of brain tumors. Multimodality management including surgery, chemotherapy, and radiation therapy remains the cornerstone in the management of pediatric brain tumors.

Expression of inducible nitric oxide synthase in carcinomas and sarcomas affecting the oral cavity.

CONTEXT: Inducible nitric oxide synthase (iNOS) is a cytoplasmic enzyme which plays a crucial role in the pathogenesis of oral carcinomas and sarcomas. AIMS: The objective of this study was to analyze the immunohistochemical expression of iNOS in carcinomas and sarcomas affecting the oral cavity in order to understand the possible role of iNOS in their biologic behavior and to correlate iNOS expression with lymph node metastasis in carcinomas and sarcomas. SETTINGS AND DESIGN: Patients, who attended the oral diagnosis department of Vinayaka Missions Sankarachariyar Dental College, were screened, for the purpose of the study. Besides these, paraffin-embedded tissue blocks were also retrieved from archives of the Oral and Maxillofacial Pathology Department. A total of 40 cases (20 carcinomas and 20 sarcomas) were selected for the study. SUBJECTS AND METHODS: A total of 40 cases (20 carcinomas and 20 sarcomas) were selected for the study. Five apparently normal tissues were obtained from the tumor adjacent normal tissue to be used as a control. These were subjected to immunohistochemical staining using antibody to iNOS and evaluated. STATISTICAL ANALYSIS USED: The results were analyzed using the Chi-square test. RESULTS: Among the 20 carcinomas 19 showed a positive immunoreactivity for iNOS and 1 case was negative. Among the 19 immunopositive iNOS cases of carcinomas, 15 cases showed positive lymph node metastasis. Among the sarcomas, positive immunoreactivity for iNOS was seen in 10 hard tissue sarcomas, while the remaining 10 soft tissue sarcomas were negative for iNOS expression. The results were analyzed using the Chi-square test. CONCLUSIONS: iNOS is a reliable marker for lymph node metastasis in carcinomas irrespective of the histologic grade. The high expression in carcinomas shows that the carcinomas elaborate more angiogenesis for growth compared with the sarcomas with the exception of hard tissue sarcomas.

Epidemiological and clinical pattern of dermatomycoses in rural India.

Superficial fungal infections are most common in tropical and subtropical countries. In this study, 297 suspected superficial fungal infection cases were identified among 15,950 patients screened. The collected samples (skin, nail, and hair) were subjected to direct microscopy with potassium hydroxide and cultured on Sabourauds dextrose agar to identify the fungal species. The prevalence of superficial fungal infection was 27.6% (82/297), dermatophytosis was 75.6% (62/82), and non-dermatophytosis was 24.4% (20/82). Among the isolated dermatophytes, Trichophyton rubrum was the commonest species (79%) and Candida (60%) the commonest non-dermatophytic species. Tinea corporis was the commonest (78%) clinical presentation.

Central odontogenic fibroma of the maxilla.

The central odontogenic fibroma (COF) is a rare benign odontogenic mesenchymal tumor of jaw bones. The World Health Organization (WHO) recognizes two variants of COF namely: 1) Epithelial-rich type (WHO) and 2) epithelial-poor type (simple type). Rare variants like ossifying COF, COF associated with giant cell lesions, and amyloid have been documented. This article presents a case of an epithelial-rich variant of COF in a 24-year-old female. It presented as a bony swelling of the maxilla and appeared as a mixed lesion in radiographs. Histopathology showed a highly cellular fibrous connective tissue stroma with plump fibroblasts and long strands of odontogenic epithelium exhibiting mild eosinophilic to clear cytoplasm. Numerous cementum-like hematoxyphilic calcifications of various sizes akin to dentin or acellular cementum were observed. We believe that clinical and radiographic features of this case may add valuable knowledge to the already existing literature.

Expression of inducible nitric oxide synthase (iNOS) in oral precancer and oral squamous cell carcinoma: an immunohistochemical study.

AIMS: Prolonged production of the free radical, nitric oxide (NO) by the enzyme inducible nitric oxide synthase (iNOS) plays an important role in tumour progression by promoting angiogenesis, invasion and inducing mutation in tumour suppressor gene. The purpose of this study is to evaluate the expression and intensity of iNOS in normal oral mucosa, precancer and oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: An immunohistochemical study was performed using rabbit monoclonal antibody to iNOS on archival formalin fixed, paraffin embedded tissues of 5 normal oral mucosal samples, 10 Leukoplakia, 10 oral submucous fibrosis (OSMF) and 15 OSCC of different grades. RESULTS: A statistical significant difference was found between normal oral mucosa, precancer and OSCC in expression of iNOS (p value 0.0015). However, there was no statistically significant difference between the expressions of iNOS within premalignant groups (p value 0.5647) and histological sub-groups of OSCC (p value-0.5647). There was no statistically significant difference in the intensity of iNOS expression within the precancer group, OSCC sub-groups and between precancer and OSCC (p value-0.623). CONCLUSION: The orderly increase in the expression of iNOS from normal, through precancer, to OSCC suggests the essential role played by iNOS in epithelial transformation and tumour formation.

A prospective, parallel group, open-labeled, comparative, multi-centric, active controlled study to evaluate the safety, tolerability and benefits of fixed dose combination of acarbose and metformin versus metformin alone in type 2 diabetes.

OBJECTIVE: The present study was a prospective, parallel group, open-labeled, comparative, multicentric, active controlled study to evaluate the safety, tolerability and benefits of fixed dose combination of acarbose and metformin versus metformin alone in type 2 diabetic patients. METHODS: A total of 229 patients with type 2 diabetes were enrolled at 5 medical centers across India. They received either acarbose (50 mg) + metformin (500 mg) bid/tid (n=115) or metformin monotherapy (500 mg) bid/ tid (n=114) for 12 weeks. Primary objective was to evaluate safety and tolerability based on the adverse events reported. Secondary objective was efficacy assessment based on changes in fasting, post prandial blood glucose and HbA1c values. RESULTS: In the acarbose + metformin group 10 patients reported 14 adverse events while in metformin group 9 patients reported 10 adverse events. No patient reported any serious adverse event or was withdraw from study because of adverse events. In the acarbose plus metformin group fasting blood glucose (FBG) decreased from a baseline of 158.85 +/- 18.14 mg/dl to 113.55 +/- 19.38 mg/dl (p < 0.0001) (decrease of 45.30 +/- 15.30 mg/dl) at 12 weeks, while in the metformin group fasting blood glucose decreased from a baseline of 158.31 +/- 26.53 mg/dl to 130.55 +/- 28.31 mg/dl (p < 0.0001) (decrease of 27.76 +/- 22.91 mg/dl) at 12 weeks. In the acarbose plus metformin group postprandial blood glucose (PPBG) decreased from a baseline of 264.65 +/- 34.03 mg/dl to 173.22 +/- 31.40 mg/dl (p < 0.0001) (decrease of 91.43 +/- 28.65 mg/dl) at 12 weeks, while in the metformin group PPBG decreased from a baseline of 253.56 +/- 36.28 mg/dl to 205.36 +/- 39.49 mg/dl (p < 0.0001) (decrease of 48.20 +/- 32.72 mg/dl) at 12 weeks. In the acarbose plus metformin group glycosylated haemoglobin (HbA1c) decreased from a baseline of 9.47 +/- 0.69% to 7.71 +/- 0.85% (p < 0.0001) (% decrease of 1.76 +/- 1.11) at 12 weeks, while in the metformin group HbAlc decreased from a baseline of 9.32 +/- 0.65% to 8.26 +/- 0.68% (p < 0.0001) (% decrease of 1.06 +/- 0.66) at 12 weeks. The combination of acarbose and metformin was found to be significantly superior in lowering the FBC (p < 0.0001), PPBG (p < 0.0001) and HbA1c (p < 0.0001) at 12 weeks as compared to metformin monotherapy. CONCLUSIONS: Fixed dose combination of acarbose and metformin was well tolerated and it was superior to metformin monotherapy in controlling FBG, PPBG and HbA(1C) levels in Type 2 Diabetes Mellitus patients.

Antigenotoxic effect of genistein against 7,12-dimethylbenz[a]anthracene induced genotoxicity in bone marrow cells of female Wistar rats.

Carcinogen induced mutation in somatic cells leads to genetic instability, which is considered as an important facet of carcinogenesis. Agents that inhibit DNA adduct formation, stimulate DNA repair mechanisms, and possess antioxidant functions are considered as antigenotoxic agents. Genistein, the major isoflavone of soy products, protects animals against experimentally induced mammary and prostate cancers. 7,12-Dimethylbenz[a]anthracene (DMBA), a potent site-specific carcinogen, induce mutations in DNA through its active metabolite, dihydrodiol epoxide, what is a crucial step in cancer initiation. The antigenotoxic effect of genistein against DMBA-induced genotoxicity has been investigated in the present study by analyzing the frequency of micronucleated polychromatic erythrocytes (MnPCEs) and chromosomal aberrations as cytogenetic end-points. The status of lipid peroxidation, antioxidants and detoxication agents were used as biochemical end-points to assess the antigenotoxic effect of genistein. Elevated MnPCEs frequency, marked chromosomal aberrations and enhanced status of lipid peroxidation, antioxidants and detoxication agents were observed in DMBA-treated animals. Oral pretreatment of genistein (20 mg/kg b.w.) for 5 days to DMBA-treated animals significantly reduced the frequency of micronucleus formation and chromosomal abnormalities as well as reversed the status of biochemical variables. Our results suggest that genistein has potent antigenotoxic effect against DMBA-induced genotoxicity.

Protective role of withaferin-A on 7,12-dimethylbenz(a)anthracene-induced genotoxicity in bone marrow of Syrian golden hamsters.

The present study has investigated the antigenotoxic effect of withaferin-A, a steroidal lactone obtained from the roots and leaves of Withania somnifera, in 7,12-dimethylbenz(a)anthracene (DMBA)-induced genotoxicity. Measurement of the frequency of micronucleated polychromatic erythrocytes (MnPCEs) and chromosomal aberrations is used as cytogenetic endpoints. A single intraperitoneal injection of DMBA (30 mg/kg b.w.) to golden Syrian hamsters resulted in marked elevation in the frequency of MnPCEs and aberrations in the chromosomal structure. Hamsters pretreated with withaferin-A intraperitonealy 2 h before the injection of DMBA, significantly reduced the frequency of MnPCEs and chromosomal aberrations such as chromosomal break, gap, minute, and fragment. Our results thus demonstrated the antigenotoxic effect of withaferin-A in DMBA-induced genotoxicity in the bone marrow of golden Syrian hamsters.

Protective effect of Withaferin-A on micronucleus frequency and detoxication agents during experimental oral carcinogenesis.

Our aim was to investigate the effect of Withaferin-A on bone marrow micronucleus frequency and buccal mucosa detoxication agents during 7, 12-dimethylbenz[a]anthracene (DMBA) induced hamster buccal pouch carcinogenesis. Oral squamous cell carcinoma was developed in hamsters' buccal pouches by painting 0.5% DMBA in liquid paraffin, three times per week for 14 weeks. We observed 100% tumor formation in DMBA painted hamsters. Elevated frequency of bone marrow micronucleated polychromatic erythrocytes (MnPCEs) and decrease in buccal mucosa phase II detoxication agents were noticed in tumor bearing hamsters. Oral administration of Withaferin-A significantly reduced the micronucleus frequency and brought back the status of phase II detoxication agents in DMBA painted hamsters. Our study thus demonstrated the protective effect of Withaferin-A on DMBA-induced micronucleus frequency in the bone marrow of golden Syrian hamsters. Also, Withaferin-A maintained the status of buccal mucosa detoxication agents during DMBA-induced hamster buccal pouch carcinogenesis.

Bacterial flavodoxins support nitric oxide production by Bacillus subtilis nitric-oxide synthase.

Unlike animal nitric-oxide synthases (NOSs), the bacterial NOS enzymes have no attached flavoprotein domain to reduce their heme and so must rely on unknown bacterial proteins for electrons. We tested the ability of two Bacillus subtilis flavodoxins (YkuN and YkuP) to support catalysis by purified B. subtilis NOS (bsNOS). When an NADPH-utilizing bacterial flavodoxin reductase (FLDR) was added to reduce YkuP or YkuN, both supported NO synthesis from either L-arginine or N-hydroxyarginine and supported a linear nitrite accumulation over a 30-min reaction period. Rates of nitrite production were directly dependent on the ratio of YkuN or YkuP to bsNOS. However, the V/Km value for YkuN (5.2 x 10(5)) was about 20 times greater than that of YkuP (2.6 x 10(4)), indicating YkuN is more efficient in supporting bsNOS catalysis. YkuN that was either photo-reduced or prereduced by FLDR transferred an electron to the bsNOS ferric heme at rates similar to those measured for heme reduction in the animal NOSs. YkuN supported a similar NO synthesis activity by a different bacterial NOS (Deinococcus radiodurans) but not by any of the three mammalian NOS oxygenase domains nor by an insect NOS oxygenase domain. Our results establish YkuN as a kinetically competent redox partner for bsNOS and suggest that FLDR/flavodoxin proteins could function physiologically to support catalysis by bacterial NOSs.

RETRACTED: Structural Stability of RNA-Binding proteins through Computation of Cation-pi Interactions.

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Structures of tryptophanyl-tRNA synthetase II from Deinococcus radiodurans bound to ATP and tryptophan. Insight into subunit cooperativity and domain motions linked to catalysis.

An auxiliary tryptophanyl tRNA synthetase (drTrpRS II) that interacts with nitric-oxide synthase in the radiation-resistant bacterium Deinococcus radiodurans charges tRNA with tryptophan and 4-nitrotryptophan, a specific nitration product of nitric-oxide synthase. Crystal structures of drTrpRS II, empty of ligands or bound to either Trp or ATP, reveal that drTrpRS II has an overall structure similar to standard bacterial TrpRSs but undergoes smaller amplitude motions of the helical tRNA anti-codon binding (TAB) domain on binding substrates. TAB domain loop conformations that more closely resemble those of human TrpRS than those of Bacillus stearothermophilus TrpRS (bsTrpRS) indicate different modes of tRNA recognition by subclasses of bacterial TrpRSs. A compact state of drTrpRS II binds ATP, from which only minimal TAB domain movement is necessary to bring nucleotide in contact with Trp. However, the signature KMSKS loop of class I synthetases does not completely engage the ATP phosphates, and the adenine ring is not well ordered in the absence of Trp. Thus, progression of the KMSKS loop to a high energy conformation that stages acyl-adenylation requires binding of both substrates. In an asymmetric drTrpRS II dimer, the closed subunit binds ATP, whereas the open subunit binds Trp. A crystallographically symmetric dimer binds no ligands. Half-site reactivity for Trp binding is confirmed by thermodynamic measurements and explained by an asymmetric shift of the dimer interface toward the occupied active site. Upon Trp binding, Asp68 propagates structural changes between subunits by switching its hydrogen bonding partner from dimer interface residue Tyr139 to active site residue Arg30. Since TrpRS IIs are resistant to inhibitors of standard TrpRSs, and pathogens contain drTrpRS II homologs, the structure of drTrpRS II provides a framework for the design of potentially useful antibiotics.

Structure and activity of an aminoacyl-tRNA synthetase that charges tRNA with nitro-tryptophan.

The most divergent of two tryptophanyl tRNA synthetases (TrpRS II) found in Deinococcus radiodurans interacts with a nitric oxide synthase protein that produces 4-nitro-tryptophan (4-NRP). TrpRS II efficiently charges transfer RNA(Trp) with 4-NRP and 5-hydroxy-tryptophan (5-HRP). The crystal structures of TrpRS II bound to tryptophan and 5-HRP reveal residue substitutions that accommodate modified indoles. A class of auxiliary bacterial TrpRSs conserve this capacity to charge tRNA with nonstandard amino acids.