Comparison of peak wall stress and peak wall rupture index in ruptured and asymptomatic intact abdominal aortic aneurysms.

BACKGROUND: Previous studies have suggested that finite element analysis (FEA) can estimate the rupture risk of an abdominal aortic aneurysm (AAA); however, the value of biomechanical estimates over measurement of AAA diameter alone remains unclear. This study aimed to compare peak wall stress (PWS) and peak wall rupture index (PWRI) in participants with ruptured and asymptomatic intact AAAs. METHODS: The reproducibility of semiautomated methods for estimating aortic PWS and PWRI from CT images was assessed. PWS and PWRI were estimated in people with ruptured AAAs and those with asymptomatic intact AAAs matched by orthogonal diameter on a 1 : 2 basis. Spearman's correlation coefficient was used to assess the association between PWS or PWRI and AAA diameter. Independent associations between PWS or PWRI and AAA rupture were identified by means of logistic regression analyses. RESULTS: Twenty individuals were included in the analysis of reproducibility. The main analysis included 50 patients with an intact AAA and 25 with a ruptured AAA. Median orthogonal diameter was similar in ruptured and intact AAAs (82·3 (i.q.r. 73·5-92·0) versus 81·0 (73·2-92·4) mm respectively; P = 0·906). Median PWS values were 286·8 (220·2-329·6) and 245·8 (215·2-302·3) kPa respectively (P = 0·192). There was no significant difference in PWRI between the two groups (P = 0·982). PWS and PWRI correlated positively with orthogonal diameter (both P < 0·001). Participants with high PWS, but not PWRI, were more likely to have a ruptured AAA after adjusting for potential confounders (odds ratio 5·84, 95 per cent c.i. 1·22 to 27·95; P = 0·027). This association was not maintained in all sensitivity analyses. CONCLUSION: High aortic PWS had an inconsistent association with greater odds of aneurysm rupture in patients with a large AAA.

Lack of an effective drug therapy for abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) rupture is a common cause of death in adults. Current AAA treatment is by open surgical or endovascular aneurysm repair. Rodent model and human epidemiology, and genetic and observational studies over the last few decades have highlighted the potential of a number of drug therapies, including medications that lower blood pressure, correct dyslipidaemia, or inhibit thrombosis, inflammation or matrix remodelling, as approaches to managing small AAA. This review summarizes prior AAA pathogenesis data from animal and human studies aimed at identifying targets for the development of drug therapies. The review also systematically assesses past randomized placebo-controlled drug trials in patients with small AAAs. Eleven previously published randomized-controlled clinical trials testing different drug therapies aimed at slowing AAA progression were identified. Five of the trials tested antibiotics and three trials assessed medications that lower blood pressure. Meta-analyses of these trials suggested that neither of these approaches limit AAA growth. Allocation to blood pressure-lowering medication was associated with a small reduction in AAA rupture or repair, compared to placebo (relative risk 0.94, 95% confidence intervals 0.89, 1.00, P = 0.047). Three further trials assessed the effect of a mast cell inhibitor, fibrate or platelet aggregation inhibition and reported no effect on AAA growth or clinical events. Past trials were noted to have a number of design issues, particularly small sample sizes and limited follow-up. Much larger trials are needed to properly test potential therapeutic approaches if a convincingly effective medical therapy for AAA is to be identified.

Studying land use dynamics using decadal satellite images and Dyna-CLUE model in the Mahanadi River basin, India.

Population growth rate indicates the proportional rate of settlement expansion and landscape modification in any river basin. The Mahanadi River basin (MRB), which is a densely populated, cropland and forest-dominated landscape, is selected as a case study area for studying the nature of built-up expansion and the corresponding land cover modifications. Satellite data-derived land use/land cover (LU/LC) maps for the years 1995, 2005, and 2015 were used for identification of landscape changes during the past three decades. One of the major LU/LC changes are observed in terms of increase in the water, which may be attributed to construction of new dams at the cost of the croplands and forest areas. Conversion of forest to cropland and expansion and densification of built-up areas in and around the existing built-up areas are also identified as a major LU/LC change. The geostatistical analysis was performed to identify the relationship between LU/LC classes with drivers, which showed that built-up areas were more in topographically flat terrain with higher soil depth, and expanded more around the existing built-up areas; cropland areas were more at lower elevation and less sloppy terrain, and forest areas were more at higher elevation. The LU/LC scenario of 2025 was projected using a spatially explicit dynamic conversion of land use and its effects (Dyna-CLUE) modeling platform with the LU/LC change trends of past 10 years (2005-2015) and 20 years (1995-2015). The major LU/LC changes observed during 2005-2015 were built-up expansion by 36.53% and deciduous forest and cropland reduction by 0.35% and 0.45%, respectively. Thus, the corresponding predicted change during 2015-2025 estimated built-up expansion by 25.70% and deciduous forest and croplands loss by 0.43% and 0.35%, respectively. On the other hand, during 1995 to 2015, the total built-up expansion and deciduous forest and cropland reduction were observed 50.79%, 0.45%, and 0.73%, respectively. Thus, the predicted changes during 2015-2025 were estimated as 18.48% built-up expansion and 0.22% and 0.21% deciduous forest and cropland loss. However, with the conditions of restricted deforestation and less landscape modification, the LU/LC projections show less built-up area expansion, reducing the cropland, fallow land, plantation, and waste land. The reduced numbers of land cover conversions types during 2005-2015 compared with 1995-2005 indicate more stabilized landscape. The input LU/LC maps and statistical analysis demonstrated the landscape modifications and causes observed in the basin. The model projected LU/LC maps are giving insights to possible changes under multiple pathways, which will help the agriculture, forest, urban, and water resource planners and managers in improved policy-making processes.

Meta-analysis of clinical trials examining the benefit of structured home exercise in patients with peripheral artery disease.

BACKGROUND: Supervised exercise is recommended for the management of peripheral artery disease (PAD); however, the uptake is limited. Structured home exercise programmes may be more feasible, but their effectiveness is unclear. This systematic review and meta-analysis examined the benefit of structured home exercise programmes for treating PAD in comparison to controls not receiving an exercise programme. METHODS: A literature search was conducted to identify RCTs comparing structured home exercise with controls not receiving an exercise programme among patients with PAD. To be included, studies had to report outcomes from treadmill or corridor walking tests, or objective assessment of physical activity. Inverse variance-weighted meta-analysis was performed to compare changes in maximum walking distance and intermittent claudication onset distance in treadmill tests, walking distance during a 6-min walking test, and physical activity measured using a pedometer or accelerometer. Summarized results are presented in terms of standard deviation differences. RESULTS: Eleven randomized trials involving 807 patients were included. Follow-up ranged from 2 to 24 months; only one trial included follow-up beyond 12 months. Meta-analyses showed that structured home exercise programmes led to significant improvements in maximum walking distance (mean difference (MD) 0·32, 95 per cent c.i. 0·15 to 0·50; P < 0·001), intermittent claudication onset distance (MD 0·45, 0·27 to 0·62; P < 0·001), walking distance in a 6-min walking test (MD 0·28, 0·09 to 0·47; P = 0·004) and physical activity (MD 0·27, 0·11 to 0·43; P = 0·001). CONCLUSION: This meta-analysis suggests that structured home exercise programmes are effective at improving walking performance and physical activity in the short term for patients with PAD.

Systematic review and meta-analysis of clinical trials examining the effect of hyperbaric oxygen therapy in people with diabetes-related lower limb ulcers.

AIM: To examine the efficacy of hyperbaric oxygen therapy in healing diabetes-related lower limb ulcers through a meta-analysis of randomized clinical trials. METHODS: A literature search was conducted to identify appropriate clinical trials. Inclusion required randomized study design and reporting of the proportion of diabetes-related lower limb ulcers that healed. A meta-analysis was performed to examine the effect of hyperbaric oxygen therapy on ulcer healing. The secondary outcomes were minor and major amputations. RESULTS: Nine randomized trials involving 585 participants were included. People allocated to hyperbaric oxygen therapy were more likely to have complete ulcer healing (relative risk 1.95, 95% CI 1.51-2.52; P<0.001), and less likely to require major (relative risk 0.54, 95% CI 0.36-0.81; P=0.003) or minor (relative risk 0.68, 95% CI 0.48-0.98; P=0.040) amputations than control groups. Sensitivity analyses suggested the findings were dependent on the inclusion of one trial. Adverse events included ear barotrauma and a seizure. Many of the trials were noted to have methodological weaknesses including absence of blinding of outcome assessors, lack of a justifiable sample size calculation and limited follow-up. CONCLUSIONS: This meta-analysis suggests hyperbaric oxygen therapy improves the healing of diabetes-related lower limb ulcers and reduces the requirement for amputation. Confidence in these results is limited by significant design weaknesses of previous trials and inconsistent findings. A more rigorous assessment of the efficacy of hyperbaric the efficacy of hyperbaric oxygen therapy is needed.

Assessment and validation of a novel angiographic scoring system for peripheral artery disease.

BACKGROUND: Angiography is used routinely in the assessment of lower-limb arteries, but there are few well validated angiographic scoring systems. The aim of this study was to develop and validate a novel angiographic scoring system for peripheral artery disease. METHODS: An angiographic scoring system (the ANGIO score) was developed and applied to a sample of patients from a single vascular surgical department who underwent CT angiography of the lower limbs. The reproducibility of the ANGIO score was compared with those of the Bollinger and Trans-Atlantic inter-Society Consensus (TASC) IIb systems in a series of randomly selected patients. Associations between the ANGIO score and lower-limb ischaemia, as measured by the ankle : brachial pressure index (ABPI), and outcome events (major lower-limb amputations and cardiovascular events - myocardial infarction, stroke and cardiovascular death) were assessed. RESULTS: Some 256 patients undergoing CT angiography were included. The interobserver reproducibility of the ANGIO score was better than that of the other scoring systems examined (κ = 0·90, P = 0·002). There was a negative correlation between the ANGIO score and ABPI (ρ = -0·33, P = 0·008). A higher ANGIO score was associated with an increased risk of major lower-limb amputation (hazard ratio (HR) for highest versus lowest tertile 9·30, 95 per cent c.i. 1·95 to 44·38; P = 0·005) and cardiovascular events (HR 2·73, 1·31 to 5·70; P = 0·007) following adjustment for established risk factors. CONCLUSION: The ANGIO score provided a reproducible and valid assessment of the severity of lower-limb ischaemia and risk of major amputation and cardiovascular events in these patients with peripheral artery disease.

Systematic Review and Meta-Analysis of the Association Between C-Reactive Protein and Major Cardiovascular Events in Patients with Peripheral Artery Disease.

BACKGROUND: Patients with peripheral artery disease (PAD) are at substantial risk of cardiovascular events. There is interest in using blood markers, such as C-reactive protein (CRP), to monitor prognosis and treatment efficacy in PAD patients. The aim of this meta-analysis was to assess the association between CRP and major cardiovascular events in PAD patients. METHOD: Studies evaluating the association between CRP and major cardiovascular events (myocardial infarction, stroke, cardiac revascularisation and mortality) were identified using MEDLINE and the Cochrane library. Studies that did not include participants with PAD, measure CRP, or follow-up patients for cardiovascular events were excluded. Meta-analyses of published adjusted hazard ratios (HR) were conducted using an inverse variance-weighted random effects model, and heterogeneity was assessed with the I2 index. RESULTS: A total of 16 studies involving 5041 participants met the inclusion criteria for the systematic review. Eight studies were included in the meta-analyses. Summary effect estimates were reported as HR comparing higher and lower quantiles, and HR per unit increase in logeCRP. PAD patients with higher CRP had a significantly greater risk of major cardiovascular events compared with those with lower CRP (HR 2.26, 95% CI 1.65-3.09, p < 0.001). The HR for major cardiovascular events was 1.38 (95% CI 1.16-1.63, p < 0.001) per unit increase in logeCRP. CONCLUSIONS: The present findings suggest that high circulating CRP is predictive of major cardiovascular events in PAD patients.

Probiotic and functional characterization of bifidobacteria of Indian human origin.

AIMS: To identify and characterize the probiotic and functional attributes of bifidobacteria isolated from human sources by in vitro methods. METHODS AND RESULTS: Twelve isolates of bifidobacteria were obtained from different human sources viz; human milk, infant and adult faeces. The preliminary identification of isolates was done using genus-specific PCR followed by species level identification using 16S rRNA sequencing. All the isolates invariably showed potential probiotic characteristics. Finally, three most promising isolates were subjected to safety evaluation and were found to be safe. These were further evaluated for their potential functional characteristics like, antioxidative, antimutagenic and cholesterol assimilation. The isolates viz; NBIF-5 (73·55 ± 0·03%) and NBIF-7 (64·06 ± 0·03%) assimilated significantly higher cholesterol than NBIF-2. The maximum antioxidative activity was observed in NBIF-2 (56·56 ± 0·28% of radical inhibition). High percentage antimutagenicity scores of 52·41 ± 2·25% and 53·68 ± 1·98% against sodium azide (NaN3) were shown by NBIF-2 and NBIF-7 respectively. CONCLUSIONS: Three strains (NBIF-2, NBIF-5 and NBIF-7) were found to be endowed with appreciable probiotic and functional activities. SIGNIFICANCE AND IMPACT OF THE STUDY: With growing demands for probioitcs and bifidobacteria being prominent ones, new strains of diversified origin have the potential to be explored for commercial and functional applications and thus can be added to the existing commercially available strains of this genera.

The shikimate pathway: review of amino acid sequence, function and three-dimensional structures of the enzymes.

The aromatic compounds such as aromatic amino acids, vitamin K and ubiquinone are important prerequisites for the metabolism of an organism. All organisms can synthesize these aromatic metabolites through shikimate pathway, except for mammals which are dependent on their diet for these compounds. The pathway converts phosphoenolpyruvate and erythrose 4-phosphate to chorismate through seven enzymatically catalyzed steps and chorismate serves as a precursor for the synthesis of variety of aromatic compounds. These enzymes have shown to play a vital role for the viability of microorganisms and thus are suggested to present attractive molecular targets for the design of novel antimicrobial drugs. This review focuses on the seven enzymes of the shikimate pathway, highlighting their primary sequences, functions and three-dimensional structures. The understanding of their active site amino acid maps, functions and three-dimensional structures will provide a framework on which the rational design of antimicrobial drugs would be based. Comparing the full length amino acid sequences and the X-ray crystal structures of these enzymes from bacteria, fungi and plant sources would contribute in designing a specific drug and/or in developing broad-spectrum compounds with efficacy against a variety of pathogens.

Antibacterial efficacy of Nisin, Pediocin 34 and Enterocin FH99 against Listeria monocytogenes and cross resistance of its bacteriocin resistant variants to common food preservatives.

Antilisterial efficiency of three bacteriocins, viz, Nisin, Pediocin 34 and Enterocin FH99 was tested individually and in combination against Listeria mononcytogenes ATCC 53135. A greater antibacterial effect was observed when the bacteriocins were combined in pairs, indicating that the use of more than one LAB bacteriocin in combination have a higher antibacterial action than when used individually. Variants of Listeria monocytogenes ATCC 53135 resistant to Nisin, Pediocin 34 and Enterocin FH99 were developed. Bacteriocin cross-resistance of wild type and their corresponding resistant variants were assessed and results showed that resistance to a bacteriocin may extend to other bacteriocins within the same class. Resistance to Pediocin 34 conferred cross resistance to Enterocin FH 99 but not to Nisin. Similarly resistance to Enterocin FH99 conferred cross resistance to Pediocin 34 but not to Nisin. Also, the sensitivity of Nisin, Pediocin 34 and Enterocin FH99 resistant variants of Listeria monocytogenes to low pH, salt, sodium nitrite, and potassium sorbate was assayed in broth and compared to the parental wild-type strain. The Nisin, Pediocin 34 and Enterocin FH99 resistant variants did not have intrinsic resistance to low pH, sodium chloride, potassium sorbate, or sodium nitrite. In no case were the bacteriocin resistant Listeria monocytogenes variants examined were more resistant to inhibitors than the parental strains.

From transabdominal to totally extra-peritoneal robotic ventral hernia repair: observations and outcomes.

PURPOSE: While robotic-assisted hernia repair has increased the popularity of minimally invasive hernia surgery, selecting between the types of approaches is a challenge for both experts and novices alike. In this study, we compared a single surgeon's early experience transitioning from transabdominal hernia repair with sublay mesh in either the pre-peritoneal or retrorectus space (TA-SM) and enhanced-view totally extra-peritoneal (eTEP) ventral hernia repair in the peri-operative and long-term post-operative time periods. METHODS: We conducted a retrospective review of 50 eTEP and 108 TA-SM procedures to collect demographics, intraoperative details, and 30-day and 1-year post-operative outcomes. Statistical analysis was performed utilizing Chi-square analysis, Fisher's test, and two sample t-tests with equal variances. RESULTS: There were no significant differences in patient demographics or comorbidities. eTEP patients had larger defects (109.1 cm2 vs. 31.8 cm2, p = 0.043) and mesh used (432.8 cm2 vs. 137.9 cm2, p = 0.001). Operative times were equivalent (158.3 ± 90.6 min eTEP and 155.8 ± 65.2 min TA-SM, p = 0.84), but conversion to alternate procedure type was higher for the transabdominal approach (4% eTEP vs. 22% TA-SM, p < 0.05). Hospital stay was less in the eTEP cohort (1.3 days vs. 2.2 days, p < 0.05). Within 30 days, there were no significant differences in emergency visits or hospital readmissions. There was a greater propensity for eTEP patients to develop seromas (12.0% vs. 1.9%, p < 0.05). At 1 year, there was no statistically significant difference in recurrence rate (4.56% eTEP vs. 12.2% TA-SM, p = 0.28) respective to average time to recurrence (9.17 months eTEP vs. 11.05 months TA-SM). CONCLUSION: The eTEP approach can be adopted safely and efficiently, and may have superior peri-operative outcomes including fewer conversions and reduced hospital stay.

Identification of a promising inhibitor from Illicium verum (star anise) against the main protease of SARS-CoV-2: insights from the computational study.

SARS-CoV-2, the causing agent of coronavirus disease (COVID-19), first broke out in Wuhan and rapidly spread worldwide, resulting in a global health emergency. The lack of specific drugs against the coronavirus has made its spread challenging to control. The main protease (Mpro) is a key enzyme of SARS-CoV-2 used as a key target in drug discovery against the coronavirus. Medicines derived from plant phytoconstituents have been widely exploited to treat various diseases. The present study has evaluated the potential of Illicium verum (star anise) phytoconstituents against Mpro by implementing a computational approach. We performed molecular docking and molecular dynamics simulation study with a set of 60 compounds to identify their potential to inhibit the main protease (Mpro) of SARS-CoV-2. DFT study and post dynamics free energy calculations were also performed to strengthen the findings. The identified four compounds by docking study exhibited the highest potential compared to other selected phytoconstituents. Further, density functional theory (DFT) calculation, molecular dynamics simulation and post dynamics MM-GBSA energy calculation predicted Verimol-G as a potential compound, which formed stable interactions through the catalytic dyad residues. The HOMO orbital energy (-0.250038) from DFT and the post dynamics binding free energy calculation (-73.33 Kcal/mol) correlate, suggesting Verimol-G is the best inhibitor compared to the other phytoconstituents. This compound also complies with the ADME properties of drug likeliness. Thus, based on a computational study, we suggest that Verimol G may be developed as a potential inhibitor against the main protease to combat COVID-19.Communicated by Ramaswamy H. Sarma.

Immunologic risk stratification of pediatric heart transplant patients by combining HLA-EMMA and PIRCHE-II.

Human leukocyte antigen (HLA) molecular mismatch is a powerful biomarker of rejection. Few studies have explored its use in assessing rejection risk in heart transplant recipients. We tested the hypothesis that a combination of HLA Epitope Mismatch Algorithm (HLA-EMMA) and Predicted Indirectly Recognizable HLA Epitopes (PIRCHE-II) algorithms can improve risk stratification of pediatric heart transplant recipients. Class I and II HLA genotyping were performed by next-generation sequencing on 274 recipient/donor pairs enrolled in the Clinical Trials in Organ Transplantation in Children (CTOTC). Using high-resolution genotypes, we performed HLA molecular mismatch analysis with HLA-EMMA and PIRCHE-II, and correlated these findings with clinical outcomes. Patients without pre-formed donor specific antibody (DSA) (n=100) were used for correlations with post-transplant DSA and antibody mediated rejection (ABMR). Risk cut-offs were determined for DSA and ABMR using both algorithms. HLA-EMMA cut-offs alone predict the risk of DSA and ABMR; however, if used in combination with PIRCHE-II, the population could be further stratified into low-, intermediate-, and high-risk groups. The combination of HLA-EMMA and PIRCHE-II enables more granular immunological risk stratification. Intermediate-risk cases, like low-risk cases, are at a lower risk of DSA and ABMR. This new way of risk evaluation may facilitate individualized immunosuppression and surveillance.

A risk-prediction model for in-hospital mortality after heart transplantation in US children.

We sought to develop and validate a quantitative risk-prediction model for predicting the risk of posttransplant in-hospital mortality in pediatric heart transplantation (HT). Children <18 years of age who underwent primary HT in the United States during 1999-2008 (n = 2707) were identified using Organ Procurement and Transplant Network data. A risk-prediction model was developed using two-thirds of the cohort (random sample), internally validated in the remaining one-third, and independently validated in a cohort of 338 children transplanted during 2009-2010. The best predictive model had four categorical variables: hemodynamic support (ECMO, ventilator support, VAD support vs. medical therapy), cardiac diagnosis (repaired congenital heart disease [CHD], unrepaired CHD vs. cardiomyopathy), renal dysfunction (severe, mild-moderate vs. normal) and total bilirubin (≥ 2.0, 0.6 to <2.0 vs. <0.6 mg/dL). The C-statistic (0.78) and the Hosmer-Lemeshow goodness-of-fit (p = 0.89) in the model-development cohort were replicated in the internal validation and independent validation cohorts (C-statistic 0.75, 0.81 and the Hosmer-Lemeshow goodness-of-fit p = 0.49, 0.53, respectively) suggesting acceptable prediction for posttransplant in-hospital mortality. We conclude that this risk-prediction model using four factors at the time of transplant has good prediction characteristics for posttransplant in-hospital mortality in children and may be useful to guide decision-making around patient listing for transplant and timing of mechanical support.

Improved survival in pediatric heart transplant recipients: have white, black and Hispanic children benefited equally?

We assessed whether the improvement in posttransplant survival in pediatric heart transplant (HT) recipients during the last two decades has benefited the major racial groups in the United States equally. We analyzed all children <18 years of age who underwent their first HT in the US during 1987-2008. We compared trends in graft loss (death or retransplant) in white, black and Hispanic children in five successive cohorts (1987-1992, 1993-1996, 1997-2000, 2001-2004, 2005-2008). The primary endpoint was early graft loss within 6 months posttransplant. Longer-term survival was assessed in recipients who survived the first 6 months. The improvement in early posttransplant survival was similar (hazard ratio [HR] for successive eras 0.80, 95% confidence interval [CI] 0.7, 0.9, p = 0.24 for black-era interaction, p = 0.22 for Hispanic-era interaction) in adjusted analysis. Longer-term survival was worse in black children (HR 2.2, CI 1.9, 2.5) and did not improve in any group with time (HR 1.0 for successive eras, CI 0.9, 1.1, p = 0.57; p = 0.19 for black-era interaction, p = 0.21 for Hispanic-era interaction). Thus, the improvement in early post-HT survival during the last two decades has benefited white, black and Hispanic children equally. Disparities in longer-term survival have not narrowed with time; the survival remains worse in black recipients.

Trends in wait-list mortality in children listed for heart transplantation in the United States: era effect across racial/ethnic groups.

We sought to evaluate trends in overall and race-specific pediatric heart transplant (HT) wait-list mortality in the United States (US) during the last 20 years. We identified all children <18 years old listed for primary HT in the US during 1989-2009 (N = 8096, 62% White, 19% Black, 13% Hispanic and 6% Other) using the Organ Procurement and Transplant Network database. Wait-list mortality was assessed in four successive eras (1989-1994, 1995-1999, 2000-2004 and 2005-2009). Overall wait-list mortality declined in successive eras (26%, 23%, 18% and 13%, respectively). The decline across eras remained significant in adjusted analysis (hazard ratio [HR] 0.70 in successive eras, 95% confidence interval [CI], 0.67-0.74) and was 67% lower for children listed during 2005-2009 versus those listed during 1989-1994 (HR 0.33; CI, 0.28-0.39). In models stratified by race, wait-list mortality decreased in all racial groups in successive eras. In models stratified by era, minority children were not at higher risk of wait-list mortality in the most recent era. We conclude that the risk of wait-list mortality among US children listed for HT has decreased by two-thirds during the last 20 years. Racial gaps in wait-list mortality present variably in the past are not present in the current era.

Biophysical Characterization of Type III Pantothenate Kinase (PanK) from Acinetobacter baumannii.

BACKGROUND: Type-III Pantothenate kinase from the multi drug resistant bacteria, Acinetobacter baumannii (AbPanK) catalyzes the first step of the essential Coenzyme A biosynthesis pathway. AbPanK is an attractive drug target against the bacteria since it is an essential enzyme and its structure is significantly different from the human PanK. METHODS: AbPanK was cloned, expressed, purified and crystallized. A good quality single crystal was used for X-ray intensity data collection. Dynamic light scattering was done for calculating the hydrodynamic radii and its oligomeric nature in the solution. Binding studies of this protein with its two substrates, Pantothenate and ATP were done using spectrofluorometer. RESULTS: Our results indicated that AbPanK shows a strong affinity with pantothenate with dissociation constant of 1.2 x 10- 8 M and moderate affinity towards ATP of 3.7x 10-3 M. This fact was further substantiated by the calculations of Km of both substrates using kinase assay kit. Dynamic light scattering studies have shown that it exists as homogenous solution with hydrodynamic radii corresponding to the molecular weight of 29.55 kDa. A low-resolution X-ray intensity data set was collected, which shows that AbPank crystallizes in P2 space group with cell dimensions of a= 165 Å, b= 260 Å, and, c= 197 Å and α= 90.0, ß= 113.60, γ= 90.0. DISCUSSION: Recombinant Pantothenate kinase from Acinetobacter baumannii was purified to homogeneity and crystallized. The enzyme exhibits very low sequence identity (28%) to other corresponding enzymes. CONCLUSION: The recombinant enzyme was active and its binding affinities with its substrates pantothenate and ATP have been studied. This information would be very useful while designing the inhibitors of this enzyme in order to fight bacterial infections associated to this pathogen.