Optimization of Enzymatic Transesterification of Acid Oil for Biodiesel Production Using a Low-Cost Lipase: The Effect of Transesterification Conditions and the Synergy of Lipases with Different Regioselectivity.

This study describes the enzymatic production of second-generation biodiesel using low-quality acid oil as a substrate. Biolipasa-R, a commercially available and low-cost lipase, was employed for enzymatic transesterification. Response surface methodology was applied to optimize the enzymatic transesterification process. The optimal conditions for biodiesel production, which comprised 42% lipase concentration (per weight of oil), 32% water content (per weight of oil), a methanol to oil molar ratio of 3:1, pH 7.0 and reaction temperature 30°C, resulted in the highest fatty acid methyl ester (FAME) content (71.3%). Subsequently, the synergistic effect of two lipases with different regioselectivities under the optimum transesterification conditions was studied, aiming at the enhancement of process efficiency. The transesterification efficiency of immobilized Biolipasa-R was determined and compared to that of Biolipasa-R in its free form. The results revealed a good performance on FAME content (66.5%), while the recycling of immobilized lipase resulted in a decrease in transesterification efficiency after three consecutive uses.

Preterm Birth Therapies to Target Inflammation.

Preterm birth (PTB; defined as delivery before 37 weeks of pregnancy) is the leading cause of morbidity and mortality in infants and children aged <5 years, conferring potentially devastating short- and long-term complications. Despite extensive research in the field, there is currently a paucity of medications available for PTB prevention and treatment. Over the past few decades, inflammation in gestational tissues has emerged at the forefront of PTB pathophysiology. Even in the absence of infection, inflammation alone can prematurely activate the main components of parturition resulting in uterine contractions, cervical ripening and dilatation, membrane rupture, and subsequent PTB. Mechanistic studies have identified critical elements of the complex inflammatory molecular pathways involved in PTB. Here, we discuss therapeutic options that target such key mediators with an aim to prevent, postpone, or treat PTB. We provide an overview of more traditional therapies that are currently used or being tested in humans, and we highlight recent advances in preclinical studies introducing novel approaches with therapeutic potential. We conclude that urgent collaborative action is required to address the unmet need of developing effective strategies to tackle the challenge of PTB and its complications.

Enzymatic Photometric Assays for the Selective Detection of Halides.

Halides are substrates and products of a number of biotechnologically important enzymes like dehalogenases, halide methyltransferases, and halogenases. Therefore, the determination of halide concentrations in samples is important. The classical methods based on mercuric thiocyanate are very dangerous, produce hazardous waste, and do not discriminate between chloride, bromide, and iodide. In this chapter, we describe a detailed protocol for the determination of halide concentrations based on the haloperoxidase-catalyzed oxidation of halides. The resulting hypohalous acids are detected using commercially available colorimetric or fluorimetric probes. The biocatalytic nature of the assays allows them to be implemented in one-pot cascade reactions with halide-generating enzymes. Since chloride is ubiquitous in biological systems, we also describe convenient photometric assays for the selective detection of bromide and iodide in the presence of chloride, obviating the need for laborious dialyses to obtain halide-free enzymes and reagents.

Recovery of Hydroxytyrosol from Olive Mill Wastewater Using the Promiscuous Hydrolase/Acyltransferase PestE.

Olive mill wastewater (OMWW) is produced annually during olive oil extraction and contains most of the health-promoting 3-hydroxytyrosol of the olive fruit. To facilitate its recovery, enzymatic transesterification of hydroxytyrosol (HT) was directly performed in an aqueous system in the presence of ethyl acetate, yielding a 3-hydroxytyrosol acetate rich extract. For this, the promiscuous acyltransferase from Pyrobaculum calidifontis VA1 (PestE) was engineered by rational design. The best mutant for the acetylation of hydroxytyrosol (PestE_I208A_L209F_N288A) was immobilized on EziG2 beads, resulting in hydroxytyrosol conversions between 82 and 89 % in one hour, for at least ten reaction cycles in a buffered hydroxytyrosol solution. Due to inhibition by other phenols in OMWW the conversions of hydroxytyrosol from this source were between 51 and 62 %. In a preparative scale reaction, 13.8 mg (57 %) of 3-hydroxytyrosol acetate was extracted from 60 mL OMWW.

Search committee diversity and applicant pool representation of women and underrepresented minorities: A quasi-experimental field study.

The diversification of applicant pools constitutes an important step for broadening the participation of women and underrepresented minorities (URMs) in the workforce. The current study focuses on recruiting diverse applicant pools in an academic setting. We test strategies grounded in homophily theory to attract a diverse set of applicants for open faculty positions. Analysis of recruitment data (13,750 job applications) showed that women search committee chairs and greater percentages of women on search committees related to more women applicants and that URMs search chairs and a greater percentage of URM members on search committees related to more URM applicants, resulting in 23% more women applicant pools with a woman chair and over 100% more URM applicants for a URM chair. Furthermore, women and URMs actively engage in ways to reach out to a more diverse set of applicants, whereas men and non-URMs' behavior maintains the status quo. We discuss the implications and advancement of homophily theory that can ultimately increase the representation of women and URM in the workforce. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Rational engineering of Luminiphilus syltensis (R)-selective amine transaminase for the acceptance of bulky substrates.

Despite the plethora of information on (S)-selective amine transaminases, the (R)-selective ones are still not well-studied; only a few structures are known to date, and their substrate scope is limited, apart from a few stellar works in the field. Herein, the structure of Luminiphilus syltensis (R)-selective amine transaminase is elucidated to facilitate engineering towards variants active on bulkier substrates. The V37A variant exhibited increased activity towards 1-phenylpropylamine and to activity against 1-butylamine. In contrast, the S248 and T249 positions, located on the ß-turn in the P-pocket, seem crucial for maintaining the activity of the enzyme.

From Natural Methylation to Versatile Alkylations Using Halide Methyltransferases.

Halide methyltransferases (HMTs) enable the enzymatic synthesis of S-adenosyl-l-methionine (SAM) from S-adenosyl-l-homocysteine (SAH) and methyl iodide. Characterisation of a range of naturally occurring HMTs and subsequent protein engineering led to HMT variants capable of synthesising ethyl, propyl, and allyl analogues of SAM. Notably, HMTs do not depend on chemical synthesis of methionine analogues, as required by methionine adenosyltransferases (MATs). However, at the moment MATs have a much broader substrate scope than the HMTs. Herein we provide an overview of the discovery and engineering of promiscuous HMTs and how these strategies will pave the way towards a toolbox of HMT variants for versatile chemo- and regioselective biocatalytic alkylations.

Directed Evolution of a Halide Methyltransferase Enables Biocatalytic Synthesis of Diverse SAM Analogs.

Biocatalytic alkylations are important reactions to obtain chemo-, regio- and stereoselectively alkylated compounds. This can be achieved using S-adenosyl-l-methionine (SAM)-dependent methyltransferases and SAM analogs. It was recently shown that a halide methyltransferase (HMT) from Chloracidobacterium thermophilum can synthesize SAM from SAH and methyl iodide. We developed an iodide-based assay for the directed evolution of an HMT from Arabidopsis thaliana and used it to identify a V140T variant that can also accept ethyl-, propyl-, and allyl iodide to produce the corresponding SAM analogs (90, 50, and 70 % conversion of 15 mg SAH). The V140T AtHMT was used in one-pot cascades with O-methyltransferases (IeOMT or COMT) to achieve the regioselective ethylation of luteolin and allylation of 3,4-dihydroxybenzaldehyde. While a cascade for the propylation of 3,4-dihydroxybenzaldehyde gave low conversion, the propyl-SAH intermediate could be confirmed by NMR spectroscopy.

Activity and specificity studies of the new thermostable esterase EstDZ2.

In this paper, we study the activity and specificity of EstDZ2, a new thermostable carboxyl esterase of unknown function, which was isolated from a metagenome library from a Russian hot spring. The biocatalytic reaction employing EstDZ2 proved to be an efficient method for the hydrolysis of aryl p-, o- or m-substituted esters of butyric acid and esters of secondary alcohols. Docking studies revealed structural features of the enzyme that led to activity differences among the different substrates.

Cervical epithelial damage promotes Ureaplasma parvum ascending infection, intrauterine inflammation and preterm birth induction in mice.

Around 40% of preterm births are attributed to ascending intrauterine infection, and Ureaplasma parvum (UP) is commonly isolated in these cases. Here we present a mouse model of ascending UP infection that resembles human disease, using vaginal inoculation combined with mild cervical injury induced by a common spermicide (Nonoxynol-9, as a surrogate for any mechanism of cervical epithelial damage). We measure bacterial load in a non-invasive manner using a luciferase-expressing UP strain, and post-mortem by qPCR and bacterial titration. Cervical exposure to Nonoxynol-9, 24 h pre-inoculation, facilitates intrauterine UP infection, upregulates pro-inflammatory cytokines, and increases preterm birth rates from 13 to 28%. Our results highlight the crucial role of the cervical epithelium as a barrier against ascending infection. In addition, we expect the mouse model will facilitate further research on the potential links between UP infection and preterm birth.

Cutinases as stereoselective catalysts: Specific activity and enantioselectivity of cutinases and lipases for menthol and its analogs.

The specific activity and enantioselectivity of immobilized cutinases from Aspergillus oryzae (AoC) and Humicola insolens (HiC) were compared with those of lipases from Thermomyces lanuginosus (TLL), Rhizomucor miehei (RML) and Lipase B from Candida antarctica (CALB) for menthol and its analogs that include isopulegol, trans-2-tert-butylcyclohexanol (2TBC), and dihydrocarveol (DHC). Common features of these alcohols are two bulky substituents: a cyclohexyl ring and an alkyl substituent. Dissimilarities are that the alkyl group reside at different positions or have dissimilar structures. The aim was to develop an understanding at a molecular level of similarities and differences in the catalytic behavior of the selected cutinases and lipases as a function of substrate structural elements. The experimental results reflect the (-)-enantioselectivity for AoC, HiC, TLL, and RML, while CALB is only active on DHC with (+)-enantioselectivity. In most cases, AoC has the highest activity while HiC is significantly more active than other enzymes on 2TBC. The E values of AoC, HiC, TLL, and RML for menthol are 27.8, 16.5, 155, and 125, respectively. HiC has a higher activity (>10-fold) on (-)-2TBC than AoC while they exhibit similar activities on menthol. Docking results reveal that the bulky group adjacent to the hydroxyl group determines the enantioselectivity of AoC, HiC, TLL, and RML. Amino acid residues that dominate the enantioselectivity of these enzymes are AoC's Phe195 aromatic ring; HiC's hydrophobic Leu 174 and Ile 169 groups; TLL's ring structures of Trp89, His258 and Tyr21; and Trp88 for RML. Results of this study highlight that cutinases can provide important advantages relative to lipases for enantioselective transformation, most notably with bulky and sterically hindered substrates.

Stress and productivity patterns of interrupted, synergistic, and antagonistic office activities.

We describe a controlled experiment, aiming to study productivity and stress effects of email interruptions and activity interactions in the modern office. The measurement set includes multimodal data for n = 63 knowledge workers who volunteered for this experiment and were randomly assigned into four groups: (G1/G2) Batch email interruptions with/without exogenous stress. (G3/G4) Continual email interruptions with/without exogenous stress. To provide context, the experiment's email treatments were surrounded by typical office tasks. The captured variables include physiological indicators of stress, measures of report writing quality and keystroke dynamics, as well as psychometric scores and biographic information detailing participants' profiles. Investigations powered by this dataset are expected to lead to personalized recommendations for handling email interruptions and a deeper understanding of synergistic and antagonistic office activities. Given the centrality of email in the modern office, and the importance of office work to people's lives and the economy, the present data have a valuable role to play.

An Empirical Study Comparing Unobtrusive Physiological Sensors for Stress Detection in Computer Work.

Several unobtrusive sensors have been tested in studies to capture physiological reactions to stress in workplace settings. Lab studies tend to focus on assessing sensors during a specific computer task, while in situ studies tend to offer a generalized view of sensors' efficacy for workplace stress monitoring, without discriminating different tasks. Given the variation in workplace computer activities, this study investigates the efficacy of unobtrusive sensors for stress measurement across a variety of tasks. We present a comparison of five physiological measurements obtained in a lab experiment, where participants completed six different computer tasks, while we measured their stress levels using a chest-band (ECG, respiration), a wristband (PPG and EDA), and an emerging thermal imaging method (perinasal perspiration). We found that thermal imaging can detect increased stress for most participants across all tasks, while wrist and chest sensors were less generalizable across tasks and participants. We summarize the costs and benefits of each sensor stream, and show how some computer use scenarios present usability and reliability challenges for stress monitoring with certain physiological sensors. We provide recommendations for researchers and system builders for measuring stress with physiological sensors during workplace computer use.

Absence of Stressful Conditions Accelerates Dexterous Skill Acquisition in Surgery.

The negative impact of strong sympathetic arousal on dexterous performance during formal surgical training is well-known. This study investigates how this relationship might change if surgical training takes place as a hobby in an informal environment. Fifteen medical students volunteered in a 5-week training regimen and weekly performed two standardized microsurgical tasks: circular cutting and simple interrupted suturing. Time was taken and two independent reviewers evaluated the surgical proficiency. The State Trait Anxiety Inventory (STAI) and the NASA Task Load Index (NASA-TLX) questionnaires measured subjective anxiety and workload, respectively. A high-resolution thermal imaging camera recorded facial imagery, from which a computational algorithm extracted the perinasal perspiration signal as indicator of sympathetic arousal. Anxiety scores on STAI questionnaires were indifferent for all five sessions. The continuously measured arousal signal from the thermal facial imagery was moderate and did not correlate with surgical proficiency or speed. Progressive experience was the strongest contributor to improved skill and speed, which were attained in record time. It appears that dexterous skill acquisition is facilitated by the absence of strong arousals, which can be naturally eliminated in the context of informal education. Given the low cost and availability of surgical simulators, this result opens the way for re-thinking the current practices in surgical training and beyond.

Scholar Plot: Design and Evaluation of an Information Interface for Faculty Research Performance.

The ability to objectively assess academic performance is critical to rewarding academic merit, charting academic policy, and promoting science. Quintessential to performing these functions is first the ability to collect valid and current data through increasingly automated online interfaces. Moreover, it is crucial to remove disciplinary and other biases from these data, presenting them in ways that support insightful analysis at various levels. Existing systems are lacking in some of these respects. Here we present Scholar Plot (SP), an interface that harvests bibliographic and research funding data from online sources. SP addresses systematic biases in the collected data through nominal and normalized metrics. Eventually, SP combines synergistically these metrics in a plot form for expert appraisal, and an iconic form for broader consumption. SP's plot and iconic forms are scalable, representing equally well individual scholars and their academic units, thus contributing to consistent ranking practices across the university organizational structure. In order to appreciate the design principles underlying SP, in particular the informativeness of nominal vs. normalized metrics, we also present the results of an evaluation survey taken by senior faculty (n = 28) with significant promotion and tenure assessment experience.

Dynamic Quantification of Migrainous Thermal Facial Patterns - A Pilot Study.

This article documents thermophysiological patterns associated with migraine episodes, where the inner canthi and supraorbital temperatures drop significantly compared to normal conditions. These temperature drops are likely due to vasoconstriction of the ophthalmic arteries under the inner canthi and sympathetic activation of the eccrine glands in the supraorbital region, respectively. The thermal patterns were observed on eight migraine patients and meticulously quantified using advance computational methods, capable of delineating small anatomical structures in thermal imagery and tracking them automatically over time. These methods open the way for monitoring migraine episodes in nonclinical environments, where the patient maintains directional attention, such as his/her computer at home or at work. This development has the potential to significantly expand the operational envelope of migraine studies.

The challenge of using isopropylamine as an amine donor in transaminase catalysed reactions.

Amine transaminases (ATAs) propose an appealing alternative to transition metal catalysts as they can provide chiral amines of high purity from pro-chiral compounds by asymmetric synthesis. Industrial interest on ATAs arises from the fact that chiral amines are present in a wide spectrum of pharmaceutical and other high value-added chiral compounds and building blocks. Despite their potential as useful synthetic tools, several drawbacks such as challenges associated with the thermodynamic equilibrium can still impede their utilization. Several methods have been developed to displace the equilibrium, such as the use of alanine as an amine donor and the subsequent removal of pyruvate with a two-enzyme system, or the use of o-xylylene diamine. To date, the preferred amine donor remains isopropylamine (IPA), as the produced acetone can be removed easily under low pressure or slight heating, without complicating the process with other enzymes. Despite its small size, IPA is not widely accepted from wild-type ATAs, and this fact compromises its wide applicability. Herein, we index the reported biocatalytic aminations with IPA, comparing the sequences, while we discuss significant parameters of the process, such as the effect of temperature and pH, as well as the protein engineering and process development advances in the field. This information is expected to provide an insight for potential designs of tailor-made ATAs and IPA processes.

Cross-disciplinary evolution of the genomics revolution.

Born out of the Human Genome Project (HGP), the field of genomics evolved with phenomenal speed into a dominant scientific and business force. While other efforts were intent on estimating the economic impact of the genomics revolution, we shift focus to the social and cultural capital generated by bridging together biology and computing-two of the constitutive disciplines of "genomics". We quantify this capital by measuring the pervasiveness of bio-computing cross-disciplinarity (XD) in genomics research during and after the HGP. To provide interlocking perspectives at the career and epistemic levels, we assembled three data sets to measure XD via (i) the collaboration network between 4190 biology and computing faculty from 155 departments in the United States, (ii) cross-departmental affiliations within a comprehensive set of human genomics publications, and (iii) the application of computational concepts and methods in research published in a preeminent genomics journal. Our results show the following: First, research featuring XD collaborations has higher citation impact than other disciplinary research-an effect observed at both the career and individual article levels. Second, genomics articles featuring XD methods tend to have higher citation impact than epistemically pure articles. Third, XD researchers of computing pedigree are drawn to the biology culture. This statistical evidence acquires deeper meaning when viewed against the organizational and knowledge transfer mechanisms revealed by the data models. With cross-disciplinary initiatives set to dominate the agenda of funding agencies, our case study provides a framework for appreciating the long-term effects of these initiatives on science and its standard-bearers.

Cloning, expression and characterization of cold active esterase (EstN7) from Bacillus cohnii strain N1: A novel member of family IV.

Esterases and lipases from extremophiles have attracted great attention due to their unique characteristics and wide applications. In the present study, an open reading frame (ORF) encoding a novel cold active esterase (EstN7) from Bacillus cohnii strain N1 was cloned and expressed in Escherichia coli. The full-length esterase gene encoding a protein of 320 amino acids with estimated molecular weight of 37.0 kDa. Amino acid sequence analysis revealed that the EstN7 belongs to family IV lipases with a characteristic penta-peptide motif (GXSXG), the catalytic triad Ser, Asp, His and the conserved HGGG motif of the family IV. The recombinant enzyme was purified to apparent homogeneity using nickel-affinity chromatography with a purification fold of 5 and recovery 94.5%. The specific activity of the purified enzyme was 336.89 U/mg. The recombinant EstN7 showed optimal activity at 5 °C moreover, EstN7 displayed full robust stability in the presence of wide range of organic solvents. The purified enzyme had Km and Vmax of 45 ±â€¯0.019 µM and 1113 µmol min-1 mg-1, respectively on p-NP-acetate. These promising characteristics of the recombinant EstN7 would underpin its possible usage with high potential in the synthesis of fragile compounds in pharmaceutical industries.