A Novel, Highly Potent NADPH-Dependent Cytochrome P450 Reductase from Waste Liza klunzingeri Liver.

The use of marine enzymes as catalysts for biotechnological applications is a topical subject. Marine enzymes usually display better operational properties than their animal, plant or bacterial counterparts, enlarging the range of possible biotechnological applications. Due to the fact that cytochrome P450 enzymes can degrade many different toxic environmental compounds, these enzymes have emerged as valuable tools in bioremediation processes. The present work describes the isolation, purification and biochemical characterization of a liver NADPH-dependent cytochrome P450 reductase (CPR) from the marine fish Liza klunzingeri (LkCPR). Experimental results revealed that LkCPR is a monomer of approximately 75 kDa that is active in a wide range of pH values (6-9) and temperatures (40-60 °C), showing the highest catalytic activity at pH 8 and 50 °C. The activation energy of the enzyme reaction was 16.3 kcal mol-1 K-1. The KM values for cytochrome C and NADPH were 8.83 µM and 7.26 µM, and the kcat values were 206.79 s-1 and 202.93 s-1, respectively. LkCPR displayed a specific activity versus cytochrome C of 402.07 µmol min-1 mg1, the highest activity value described for a CPR up to date (3.2-4.7 times higher than the most active reported CPRs) and showed the highest thermostability described for a CPR. Taking into account all these remarkable catalytic features, LkCPR offers great potential to be used as a suitable biocatalyst.

Biocatalysis: An Eco-Friendly Scenario for the Manufacturing of APIs.

Nowadays, the worldwide demand for Active Pharmaceutical Ingredients (APIs) requires novel, cost-effective, safe, and environmentally friendly synthetic processes [...].

Impact of different COVID-19 waves on kidney replacement therapy epidemiology and mortality: REMER 2020.

BACKGROUND: Kidney replacement therapy (KRT) confers the highest risk of death from coronavirus disease 2019 (COVID-19). However, most data refer to the early pandemic waves. Whole-year analysis compared with prior secular trends are scarce. METHODS: We present the 2020 REMER Madrid KRT registry, corresponding to the Spanish Region hardest hit by COVID-19. RESULTS: In 2020, KRT incidence decreased 12% versus 2019, while KRT prevalence decreased by 1.75% for the first time since records began and the number of kidney transplants (KTs) decreased by 16%. Mortality on KRT was 10.2% (34% higher than the mean for 2008-2019). The 2019-2020 increase in mortality was larger for KTs (+68%) than for haemodialysis (+24%) or peritoneal dialysis (+38%). The most common cause of death was infection [n = 419 (48% of deaths)], followed by cardiovascular [n = 200 (23%)]. Deaths from infection increased by 167% year over year and accounted for 95% of excess deaths in 2020 over 2019. COVID-19 was the most common cause of death (68% of infection deaths, 33% of total deaths). The bulk of COVID-19 deaths [209/285 (73%)] occurred during the first COVID-19 wave, which roughly accounted for the increased mortality in 2020. Being a KT recipient was an independent risk factor for COVID-19 death. CONCLUSIONS: COVID-19 negatively impacted the incidence and prevalence of KRT, but the increase in KRT deaths was localized to the first wave of the pandemic. The increased annual mortality argues against COVID-19 accelerating the death of patients with short life expectancy and the temporal pattern of COVID-19 mortality suggests that appropriate healthcare may improve outcomes.

Rational Design of a Thermostable 2'-Deoxyribosyltransferase for Nelarabine Production by Prediction of Disulfide Bond Engineering Sites.

One of the major drawbacks of the industrial implementation of enzymatic processes is the low operational stability of the enzymes under tough industrial conditions. In this respect, the use of thermostable enzymes in the industry is gaining ground during the last decades. Herein, we report a structure-guided approach for the development of novel and thermostable 2'-deoxyribosyltransferases (NDTs) based on the computational design of disulfide bonds on hot spot positions. To this end, a small library of NDT variants from Lactobacillus delbrueckii (LdNDT) with introduced cysteine pairs was created. Among them, LdNDTS104C (100% retained activity) was chosen as the most thermostable variant, displaying a six- and two-fold enhanced long-term stability when stored at 55 °C (t1/255 °C ≈ 24 h) and 60 °C (t1/260 °C ≈ 4 h), respectively. Moreover, the biochemical characterization revealed that LdNDTS104C showed >60% relative activity across a broad range of temperature (30−90 °C) and pH (5−7). Finally, to study the potential application of LdNDTS104C as an industrial catalyst, the enzymatic synthesis of nelarabine was successfully carried out under different substrate conditions (1:1 and 3:1) at different reaction times. Under these experimental conditions, the production of nelarabine was increased up to 2.8-fold (72% conversion) compared with wild-type LdNDT.

Magnetic Multi-Enzymatic System for Cladribine Manufacturing.

Enzyme-mediated processes have proven to be a valuable and sustainable alternative to traditional chemical methods. In this regard, the use of multi-enzymatic systems enables the realization of complex synthetic schemes, while also introducing a number of additional advantages, including the conversion of reversible reactions into irreversible processes, the partial or complete elimination of product inhibition problems, and the minimization of undesirable by-products. In addition, the immobilization of biocatalysts on magnetic supports allows for easy reusability and streamlines the downstream process. Herein we have developed a cascade system for cladribine synthesis based on the sequential action of two magnetic biocatalysts. For that purpose, purine 2'-deoxyribosyltransferase from Leishmania mexicana (LmPDT) and Escherichia coli hypoxanthine phosphoribosyltransferase (EcHPRT) were immobilized onto Ni2+-prechelated magnetic microspheres (MagReSyn®NTA). Among the resulting derivatives, MLmPDT3 (activity: 11,935 IU/gsupport, 63% retained activity, operational conditions: 40 °C and pH 5-7) and MEcHPRT3 (12,840 IU/gsupport, 45% retained activity, operational conditions: pH 5-8 and 40-60 °C) emerge as optimal catalysts for further synthetic application. Moreover, the MLmPDT3/MEcHPRT3 system was biochemically characterized and successfully applied to the one-pot synthesis of cladribine under various conditions. This methodology not only displayed a 1.67-fold improvement in cladribine synthesis (compared to MLmPDT3), but it also implied a practically complete transformation of the undesired by-product into a high-added-value product (90% conversion of Hyp into IMP). Finally, MLmPDT3/MEcHPRT3 was reused for 16 cycles, which displayed a 75% retained activity.

The Impact of Immunosuppression and Autoimmune Disease on Severe Outcomes in Patients Hospitalized with COVID-19.

Immunosuppression (IS) and autoimmune disease (AD) are prevalent in patients with severe coronavirus disease 2019 (COVID-19), but their impact on its clinical course is unknown. We investigated relationships between IS, AD, and outcomes in patients hospitalized with COVID-19. Data on consecutive admissions for COVID-19 were extracted retrospectively from medical records. Patients were assigned to one of four cohorts, according to whether or not they had an AD (AD and NAD) or were immunosuppressed (IS and NIS). The primary endpoint was development of severe acute respiratory distress syndrome (ARDS); secondary endpoints included death, and a composite of mechanical ventilation (MV) or death. A total of 789 patients were included: 569 (72.1%) male, 76 (9.6%) with an AD, and 63 (8.0%) with IS. Relative to the NIS-NAD cohort, patients in the IS-AD cohort had a significantly reduced risk of severe ARDS (adjusted hazard ratio [aHR] 0.42; 95% confidence interval [CI] 0.23-0.80; p = 0.008). No significant relationships between IS or AD status and either death or the composite of MV and death were identified, although a trend towards higher mortality was identified in the IS-NAD cohort (aHR vs NIS-NAD 1.71; 95% CI 0.94-3.12; p = 0.081). Patients in this cohort also had higher median serum levels of interleukin-6 compared with IS-AD patients (98.2 vs 21.6 pg/mL; p = 0.0328) and NIS-NAD patients (29.1 pg/mL; p = 0.0057). In conclusion, among patients hospitalized with COVID-19, those receiving immunosuppressive treatment for an AD may have a reduced risk of developing severe ARDS.

Absolute blood volume variations and vascular refilling in hemodialysis patients.

The imbalance between ultrafiltration volume (UF) and vascular refilling is considered a major cause for intradialytic hypotension. Recent studies report a noninvasive method to estimate vascular refilling (VREF ) by determining absolute blood volume (ABV). It was the aim of the study to analyze variations in ABV in a group of hemodialysis (HD) patients and examine VREF . Thirty one stable chronic HD patients were studied, aged 71.07 ± 13.31 years. Dialysis duration and UF requirements were based on physician prescription. VREF was calculated as: VREF  = VUF  - ΔV where ΔV is ABV variation during dialysis treatment. ABV at the beginning of the dialysis was 6.00 ± 2.39 L (92.82 ± 33.17 ml/kg) and at the end 5.38 ± 2.32 L (82.07 ± 31.41 ml/kg). Prescribed UF was 2.64 ± 0.83 L. Mean VREF was 2.05 ± 0.80 L, with a refilling fraction of 75.75 ± 12.79%. VREF was strongly correlated with UF volume (r2 0.877), and with pre-dialysis volume overload (r2 0.617). Patients under beta-blocker treatment showed significantly lower FREF . ABV measurement is an easy and noninvasive method that allows us to study VREF during HD. We found a strong correlation between VREF and UF.

Absolute blood volume variations during hemodialysis: Does dialysate temperature play a role?

BACKGROUND: Vascular refilling occurs to preserve hemodynamic stability during hemodialysis (HD). Recent studies report a feasible and noninvasive method to determine absolute blood volume (ABV), and estimate vascular refilling during HD. The objective of this study is to analyze if lowering dialysate temperature modifies variations in ABV during HD. METHODS: The study was performed in 50 patients under HD. During two different sessions, relative blood volume was assessed using dialysate temperatures of 35.5°C (cool dialysate) and 36.5°C (neutral dialysate). ABV and vascular refilling were calculated using Kron et al methodology. RESULTS: Thirty-nine intradialytic morbid events (IMEs) were observed in 30 patients, 14 under cool dialysate and 25 during neutral dialysate. We did not found statistically differences in ABV or in refilling volume between cool and neutral temperature. When analyzing apart only those patients who presented IME, we observed lower drop in ABV in the 35.5°C dialysate treatments (0.57 L) versus 36.5°C dialysate treatments (0.71 L). When cool dialysate was used, the vascular refilling fraction tended to be higher, but data did not turn statistically significant. CONCLUSIONS: In selected groups of patients the use of cool dialysate induces lower ABV variations that could improve hemodynamic stability during HD treatments.

Peritoneal dialysis (PD) technique training: what features influence learning time?

BACKGROUND: Within peritoneal dialysis (PD) complications, peritonitis remains a primary challenge for the long-term success of the technique. Proper technique training is essential, since it reduces peritonitis rates, but the adequacy of training has not been standardized. Furthermore, factors influencing training duration have not been well identified. METHODS: We retrospectively analyzed all consecutive training sessions of incident PD patients in our Unit from January 2001 to December 2018. RESULTS: Our analysis included 135 patients, 25.9% were diabetic and median Charlson index (CCI) was 4 (IQR 2-6). Above 13 sessions was chosen as the cut off between usual and prolonged training, as it was our cohort's 75th percentile: 23% (31) had an extended training duration as per our study definition and 77% (104) had a usual training duration. The number of training sessions required increased with age (Spearman Rho 0.404; p = 0.000001), diabetic status (p = 0.001), unemployment status (p = 0.046) and CCI (Spearman Rho 0.369; p = 0.00001). Neither gender, cohabitation status, scheduled PD start, education level nor referral origin, were significant factors impacting training duration. Requiring longer training (> 13 sessions) was a significant risk factor for higher peritonitis risk, but extended training was not related to a shorter technique survival. CONCLUSION: Number of PD training sessions depends on the patient's age and comorbidities, but is not related to social, educational or employment status. Prolonged training duration was a statistically significant predictor of higher peritonitis risk, but it was not related to shorter permanence in PD in our series. Identifying these patients since the training period would be useful to adapt training schedule as an early prevention strategy to minimize the risk of peritonitis and plan a preemptive retraining.

Ventilator Mode Does Not Influence Blood Loss or Transfusion Requirements During Major Spine Surgery: A Retrospective Study.

BACKGROUND: Blood loss during adult spinal deformity surgery is multifactorial. Anesthetic-related factors, such as mode of mechanical ventilation, may contribute to intraoperative blood loss. The aim of this study was to determine the influence of ventilator mode and ventilator parameters on intraoperative blood loss and transfusion requirements in patients undergoing prone position spine surgery. METHODS: This single-center retrospective study examined electronic medical records of patients ≥18 years of age who underwent elective prone position spine surgery between May 2015 and June 2016. Associations between ventilator mode and ventilator parameters with intraoperative estimated blood loss (EBL), packed red blood cells (PRBCs), fresh-frozen plasma (FFP), cryoprecipitate and platelet transfusions, and subfascial drain output were examined using multiple linear regression models controlling for age, sex, American Society of Anesthesiologist (ASA) physical status score, body mass index (BMI), preoperative blood coagulation parameters and laboratory values, operative levels, cage constructs, osteotomies, transforaminal lumbar interbody fusions, laminectomies, reoperation, spine surgery invasiveness index, and operative time. In a secondary analysis, EBL, blood product transfusions, and postoperative drain output were compared between pressure-controlled ventilation (PCV) and volume-controlled ventilation (VCV) propensity score-matched cohorts. RESULTS: Nine hundred forty-six records were reviewed, and 822 were included in the analysis. After adjusting for confounding, no statistically significant associations were observed between mode of ventilation and intraoperative EBL (estimate, -2; 95% confidence interval [CI], -248 to 245; P = .99) or blood product transfusions (PRBC: estimate, -9; 95% CI, -154 to 135; P = .90; FFP: estimate, -3; 95% CI, -59 to 54; P = .93; cryoprecipitate: estimate, -14; 95% CI, -70 to 43; P = .63; platelets: -7; 95% CI, -39 to 24; P = .64). After propensity score matching (n = 27 per group), no significant differences were observed in EBL (mean difference, 525 mL; 95% CI, -15 to 1065; P = .056) or blood transfusions (PRBC: mean difference, 208 mL; 95% CI, -23 to 439; P = .077; FFP (mean difference, 34 mL; 95% CI, -17 to 84; P = .19); cryoprecipitate (mean difference, 55 mL; 95% CI, -24 to 133; P = .17); or platelets (mean difference, 26 mL; 95% CI, -12 to 64; P = .18) between PCV and VCV groups. CONCLUSIONS: In prone position spine surgery, neither mode of mechanical ventilation nor airway pressure is associated with intraoperative blood loss or need for allogeneic transfusion. Use of modern ventilation strategies using lung protective techniques may mitigate differences in blood loss previously observed between PCV and VCV modes.

Resuming peritoneal dialysis after catheter removal due to complicated peritonitis.

BACKGROUND: Causes of non-resuming peritoneal dialysis (PD) after complicated peritonitis requiring peritoneal catheter (PC) removal remain poorly studied. METHODS: We reviewed all peritonitis episodes in our center between 1997 and 2017. Patients who restarted PD after PC removal (Group 1) were compared to those who did not (Group 2), identifying the causes. RESULTS: Of 284 peritonitis episodes, PC was removed in 48 patients (16.9%). In 18 (37.5%) patients PC was reinserted, and PD successfully resumed in all, with a median duration of PD afterwards of 14.1 months. In other 30 (62.5%) reinsertion of PC was not attempted. Causes of non-reinsertion were: transfer to hemodialysis 76.6% (n = 23), death 16.7% (n = 5) and transplantation 6.7% (n = 2). Hemodialysis switch was due to non-medical reasons in 47.8% (n = 11) including fear of peritonitis, family decision and social dependence. Group 1 was younger (p = 0.041), with lower Charlson index (p = 0.045) and higher men proportion (p = 0.049). Group 1 had a better patient survival than group 2 (survival at 24 months: 67% and 53%, respectively; log-rank test p: 0.01). There were no differences in survival between groups when adjusted for significant basal characteristics. CONCLUSIONS: Resuming PD after severe peritonitis requiring PC removal is feasible but a high proportion of patients do not restart PD for non-medical reasons, usually older patients with higher Charlson index. A properly structured interview would be a useful tool that could improve return to technique in these patients.

Structure-Guided Tuning of a Selectivity Switch towards Ribonucleosides in Trypanosoma brucei Purine Nucleoside 2'-Deoxyribosyltransferase.

The use of nucleoside 2'-deoxyribosyltransferases (NDTs) as biocatalysts for the industrial synthesis of nucleoside analogues is often hindered by their strict preference for 2'-deoxyribonucleosides. It is shown herein that a highly versatile purine NDT from Trypanosoma brucei (TbPDT) can also accept ribonucleosides as substrates; this is most likely because of the distinct role played by Asn53 at a position that is usually occupied by Asp in other NDTs. Moreover, this unusual activity was improved about threefold by introducing a single amino acid replacement at position 5, following a structure-guided approach. Biophysical and biochemical characterization revealed that the TbPDTY5F variant is a homodimer that displays maximum activity at 50 °C and pH 6.5 and shows a remarkably high melting temperature of 69 °C. Substrate specificity studies demonstrate that 6-oxopurine ribonucleosides are the best donors (inosine>guanosine≫adenosine), whereas no significant preferences exist between 6-aminopurines and 6-oxopurines as base acceptors. In contrast, no transferase activity could be detected on xanthine and 7-deazapurines. TbPDTY5F was successfully employed in the synthesis of a wide range of modified ribonucleosides containing different purine analogues.

Reaction mechanism of nucleoside 2'-deoxyribosyltransferases: free-energy landscape supports an oxocarbenium ion as the reaction intermediate.

Insight into the catalytic mechanism of Lactobacillus leichmannii nucleoside 2'-deoxyribosyltransferase (LlNDT) has been gained by calculating a quantum mechanics-molecular mechanics (QM/MM) free-energy landscape of the reaction within the enzyme active site. Our results support an oxocarbenium species as the reaction intermediate and thus an SN1 reaction mechanism in this family of bacterial enzymes. Our mechanistic proposal is validated by comparing experimental kinetic data on the impact of the single amino acid replacements Tyr7, Glu98 and Met125 with Ala, Asp and Ala/norLeu, respectively, and accounts for the specificity shown by this enzyme on a non-natural substrate. This work broadens our understanding of enzymatic C-N bond cleavage and C-N bond formation.

Comparison of Coagulation Parameters in Arterial and Venous Blood in Cardiac Surgery Measured Using the Quantra System.

OBJECTIVE: Perioperative coagulation testing often is performed with arterial samples even though device reference ranges typically are established in venous samples. Although limited studies exist comparing coagulation parameters across sampling sites, viscoelastic testing devices have demonstrated some differences. The objective of this study was to compare coagulation parameters determined using the Quantra System for venous and arterial samples. DESIGN: Prospective, observational study. SETTING: Tertiary care university hospital. PARTICIPANTS: The study comprised 30 adult patients undergoing cardiac surgery. INTERVENTIONS: Paired arterial and venous samples were obtained at 2 of the following time points: baseline, during bypass, or after protamine reversal of heparin. Quantra measurements included Clot Time (CT), Heparinase Clot Time (CTH), Clot Time Ratio (CTR), Clot Stiffness (CS), and Fibrinogen (FCS) and Platelet (PCS) Contributions to clot stiffness. MEASUREMENTS AND MAIN RESULTS: The relationship and agreement between matched data pairs were established and statistical analysis was performed via paired t tests. CTR, CS, FCS, and PCS were unaffected by the sampling site, whereas CT and CTH demonstrated statistically significant differences between arterial and venous samples (p < 0.001). Arterial clot times were prolonged relative to the venous ones with a mean percent error of 14.2 % and 11.9 %, respectively. These results are in general agreement with those reported for other viscoelastic testing devices. CONCLUSIONS: This study demonstrates that Quantra clot stiffness-based parameters (CS, FCS, PCS) are unaffected by sampling site, whereas the clot time parameters (CT and CTH) show good correlation in the presence of a bias. CTR, a ratio of CT and CTH, also is unaffected.

Purine and pyrimidine salvage pathway in thermophiles: a valuable source of biocatalysts for the industrial production of nucleic acid derivatives.

Due to their similarity to natural counterparts, nucleic acid derivatives (nucleobases, nucleosides, and nucleotides, among others) are interesting molecules for pharmaceutical, biomedical, or food industries. For this reason, there is increasing worldwide demand for the development of efficient synthetic processes for these compounds. Chemical synthetic methodologies require numerous protection-deprotection steps and often lead to the presence of undesirable by-products or enantiomeric mixtures. These methods also require harsh operating conditions, such as the use of organic solvents and hazard reagents. Conversely, enzymatic production by whole cells or enzymes improves regio-, stereo-, and enantioselectivity and provides an eco-friendly alternative. Because of their essential role in purine and pyrimidine scavenging, enzymes from purine and pyrimidine salvage pathways are valuable candidates for the synthesis of many different nucleic acid components. In recent years, many different enzymes from these routes, such as nucleoside phosphorylases, nucleoside kinases, 2'-deoxyribosyltransferases, phosphoribosyl transferases, or deaminases, have been successfully employed as biocatalysts in the production of nucleobase, nucleoside, or nucleotide analogs. Due to their great activity and stability at extremely high temperatures, the use of enzymes from thermophiles in industrial biocatalysis is gaining momentum. Thermophilic enzymes not only display unique characteristics such as temperature, chemical, and pH stability but also provide many different advantages from an industrial perspective. This mini-review aims to cover the most representative enzymatic approaches for the synthesis of nucleic acid derivatives. In this regard, we provide detailed comments about enzymes involved in crucial steps of purine and pyrimidine salvage pathways in thermophiles, as well as their biological role, biochemical characterization, active site mechanism, and substrate specificity. In addition, the most interesting synthetic examples reported in the literature are also included.

Marine chitinolytic enzymes, a biotechnological treasure hidden in the ocean?

Chitinolytic enzymes are capable to catalyze the chitin hydrolysis. Due to their biomedical and biotechnological applications, nowadays chitinolytic enzymes have attracted worldwide attention. Chitinolytic enzymes have provided numerous useful materials in many different industries, such as food, pharmaceutical, cosmetic, or biomedical industry. Marine enzymes are commonly employed in industry because they display better operational properties than animal, plant, or bacterial homologs. In this mini-review, we want to describe marine chitinolytic enzymes as versatile enzymes in different biotechnological fields. In this regard, interesting comments about their biological role, reaction mechanism, production, functional characterization, immobilization, and biotechnological application are shown in this work.

Characterization of an atypical, thermostable, organic solvent- and acid-tolerant 2'-deoxyribosyltransferase from Chroococcidiopsis thermalis.

In our search for thermophilic and acid-tolerant nucleoside 2'-deoxyribosyltransferases (NDTs), we found a good candidate in an enzyme encoded by Chroococcidiopsis thermalis PCC 7203 (CtNDT). Biophysical and biochemical characterization revealed CtNDT as a homotetramer endowed with good activity and stability at both high temperatures (50-100 °C) and a wide range of pH values (from 3 to 7). CtNDT recognizes purine bases and their corresponding 2'-deoxynucleosides but is also proficient using cytosine and 2'-deoxycytidine as substrates. These unusual features preclude the strict classification of CtNDT as either a type I or a type II NDT and further suggest that this simple subdivision may need to be updated in the future. Our findings also hint at a possible link between oligomeric state and NDT's substrate specificity. Interestingly from a practical perspective, CtNDT displays high activity (80-100%) in the presence of several water-miscible co-solvents in a proportion of up to 20% and was successfully employed in the enzymatic production of several therapeutic nucleosides such as didanosine, vidarabine, and cytarabine.

Effectiveness of Psycho-Physiological Portable Devices to Analyse Effect of Ergogenic Aids in Military Population.

Caffeine is one of the ergogenic substances most used by warfighters in current operation areas, but the effect on the organic response and operational performed is already poor knowledge. This research aimed to study the acute effect of 400 mg of caffeine monohydrate on the psycho-physiological response and rifle marksmanship of warfighters during a close quarter combat simulation. We analysed parameter of heart rate, blood lactate, cortical arousal, state anxiety and marksmanship of 20 Spanish Army veteran warfighters before and after a close quarter combat simulation in a double-blind procedure, also a correlation analysis was conducted between all the study variables. Marksmanship of warfighters did not improve with the caffeine ingestion, but it produced an increase in blood lactate concentration (caffeine: 1.9 ± 0.5 vs. 9.2 ± 1.1 mmol.l-1; placebo: 1.8 ± 0.7 vs. 6.9 ± 2.2 mmol.l-1), cortical arousal (% of change: caffeine: 2.51; placebo: -1.96) and heart rate (caffeine: 80.0 ± 7.2 vs. 172.9 ± 28.2 bpm; placebo: 79.3 ± 6.4 vs. 154.0 ± 26.8 bpm). In addition, higher heart rate values correlated negatively with marksmanship in close quarter combat. We concluded that caffeine intake did not improve the warfighters rifle marksmanship in close quarter combat possibly due to the increase in the physiological response.

2'-Deoxyribosyltransferase from Leishmania mexicana, an efficient biocatalyst for one-pot, one-step synthesis of nucleosides from poorly soluble purine bases.

Processes catalyzed by enzymes offer numerous advantages over chemical methods although in many occasions the stability of the biocatalysts becomes a serious concern. Traditionally, synthesis of nucleosides using poorly water-soluble purine bases, such as guanine, xanthine, or hypoxanthine, requires alkaline pH and/or high temperatures in order to solubilize the substrate. In this work, we demonstrate that the 2'-deoxyribosyltransferase from Leishmania mexicana (LmPDT) exhibits an unusually high activity and stability under alkaline conditions (pH 8-10) across a broad range of temperatures (30-70 °C) and ionic strengths (0-500 mM NaCl). Conversely, analysis of the crystal structure of LmPDT together with comparisons with hexameric, bacterial homologues revealed the importance of the relationships between the oligomeric state and the active site architecture within this family of enzymes. Moreover, molecular dynamics and docking approaches provided structural insights into the substrate-binding mode. Biochemical characterization of LmPDT identifies the enzyme as a type I NDT (PDT), exhibiting excellent activity, with specific activity values 100- and 4000-fold higher than the ones reported for other PDTs. Interestingly, LmPDT remained stable during 36 h at different pH values at 40 °C. In order to explore the potential of LmPDT as an industrial biocatalyst, enzymatic production of several natural and non-natural therapeutic nucleosides, such as vidarabine (ara A), didanosine (ddI), ddG, or 2'-fluoro-2'-deoxyguanosine, was carried out using poorly water-soluble purines. Noteworthy, this is the first time that the enzymatic synthesis of 2'-fluoro-2'-deoxyguanosine, ara G, and ara H by a 2'-deoxyribosyltransferase is reported.

Psycho-physiological response in an automatic parachute jump.

Parachute jump is an extreme activity that elicits an intense stress response that affects jumpers' body systems being able to put them at risk. The present research analysed modifications in blood oxygen saturation (BOS), heart rate (HR), cortisol, glucose, lactate, creatine kinase (CK), muscles strength, cortical arousal, autonomic modulation, pistol magazine reload time (PMRT) and state anxiety before and after an automatic open parachute jump in 38 male Spanish soldiers (25.6 ± 5.9 years). A MANOVA with samples as a fixed factor and Effect Size (ES) were conducted. MANOVA showed (Wilks lambda = .225; F = 5.980; P = .000) a significantly increase in cortisol (6.2 ± 3.2 vs. 8.2 ± 4.3 nmol/l; P = .025; ES = .47), HR (75.0 ± 14.6 vs. 87.4 ± 17.3 bpm; P = .004; ES = .72), lactate (1.8 ± 1.2 vs. 4.4 ± 2.2 mmol · l-1; P = .002; ES = 1.18), sympathetic nervous system and leg strength manifestation after the parachute jump. By contrary BOS, PMRT (55.6 ± 27.6 vs. 48.0 ± 16.7 s; P = .021; ES = .46) and somatic anxiety (SA), evaluated by CSAI2R questionnaire, decreased. An automatic parachute jump increased physiological and cortical response and decreased SA of participants. This stress response can affect the jumpers' abilities and allow us to have a better understanding of the organism stress response and to improve training for both military and sport parachute jumps.