scholarly journals Filling the Void: Introducing Aromatic Interactions into Solvent Tunnels To Enhance Lipase Stability in Methanol

2018 ◽  
Vol 84 (23) ◽  
Author(s):  
Shalev Gihaz ◽  
Margarita Kanteev ◽  
Yael Pazy ◽  
Ayelet Fishman
Keyword(s):  
Organic Solvents ◽  
Inner Core ◽  
Intramolecular Interactions ◽  
Rational Approach ◽  
Geobacillus Stearothermophilus ◽  
Content Type ◽  
Aromatic Residues ◽  
Aromatic Interactions ◽  
Biodiesel Synthesis ◽  
Enhanced Stability

ABSTRACT An enhanced stability of enzymes in organic solvents is desirable under industrial conditions. The potential of lipases as biocatalysts is mainly limited by their denaturation in polar alcohols. In this study, we focused on selected solvent tunnels in lipase from Geobacillus stearothermophilus T6 to improve its stability in methanol during biodiesel synthesis. Using rational mutagenesis, bulky aromatic residues were incorporated to occupy solvent channels and induce aromatic interactions leading to a better inner core packing. The chemical and structural characteristics of each solvent tunnel were systematically analyzed. Selected residues were replaced with Phe, Tyr, or Trp. Overall, 16 mutants were generated and screened in 60% methanol, from which 3 variants showed an enhanced stability up to 81-fold compared with that of the wild type. All stabilizing mutations were found in the longest tunnel detected in the “closed-lid” X-ray structure. The combination of Phe substitutions in an A187F/L360F double mutant resulted in an increase in unfolding temperature (Tm) of 7°C in methanol and a 3-fold increase in biodiesel synthesis yield from waste chicken oil. A kinetic analysis with p-nitrophenyl laurate revealed that all mutants displayed lower hydrolysis rates (kcat), though their stability properties mostly determined the transesterification capability. Seven crystal structures of different variants were solved, disclosing new π-π or CH/π intramolecular interactions and emphasizing the significance of aromatic interactions for improved solvent stability. This rational approach could be implemented for the stabilization of other enzymes in organic solvents. IMPORTANCE Enzymatic synthesis in organic solvents holds increasing industrial opportunities in many fields; however, one major obstacle is the limited stability of biocatalysts in such a denaturing environment. Aromatic interactions play a major role in protein folding and stability, and we were inspired by this to redesign enzyme voids. The rational protein engineering of solvent tunnels of lipase from Geobacillus stearothermophilus is presented here, offering a promising approach to introduce new aromatic interactions within the enzyme core. We discovered that longer tunnels leading from the surface to the enzyme active site were more beneficial targets for mutagenesis for improving lipase stability in methanol during biodiesel biosynthesis. A structural analysis of the variants confirmed the generation of new interactions involving aromatic residues. This work provides insights into stability-driven enzyme design by targeting the solvent channel void.

scholarly journals Protein Engineering by Random Mutagenesis and Structure-Guided Consensus of Geobacillus stearothermophilus Lipase T6 for Enhanced Stability in Methanol

2013 ◽  
Vol 80 (4) ◽  
pp. 1515-1527 ◽  
Author(s):  
Adi Dror ◽  
Einav Shemesh ◽  
Natali Dayan ◽  
Ayelet Fishman
Keyword(s):  
Protein Engineering ◽  
Organic Solvents ◽  
Random Mutagenesis ◽  
Acid Methyl Ester ◽  
Biodiesel Production ◽  
Geobacillus Stearothermophilus ◽  
Screening Assay ◽  
Lipase Gene ◽  
Improved Stability ◽  
Enhanced Stability

ABSTRACTThe abilities of enzymes to catalyze reactions in nonnatural environments of organic solvents have opened new opportunities for enzyme-based industrial processes. However, the main drawback of such processes is that most enzymes have a limited stability in polar organic solvents. In this study, we employed protein engineering methods to generate a lipase for enhanced stability in methanol, which is important for biodiesel production. Two protein engineering approaches, random mutagenesis (error-prone PCR) and structure-guided consensus, were applied in parallel on an unexplored lipase gene fromGeobacillus stearothermophilusT6. A high-throughput colorimetric screening assay was used to evaluate lipase activity after an incubation period in high methanol concentrations. Both protein engineering approaches were successful in producing variants with elevated half-life values in 70% methanol. The best variant of the random mutagenesis library, Q185L, exhibited 23-fold-improved stability, yet its methanolysis activity was decreased by one-half compared to the wild type. The best variant from the consensus library, H86Y/A269T, exhibited 66-fold-improved stability in methanol along with elevated thermostability (+4.3°C) and a 2-fold-higher fatty acid methyl ester yield from soybean oil. Based onin silicomodeling, we suggest that the Q185L substitution facilitates a closed lid conformation that limits access for both the methanol and substrate excess into the active site. The enhanced stability of H86Y/A269T was a result of formation of new hydrogen bonds. These improved characteristics make this variant a potential biocatalyst for biodiesel production.

scholarly journals Genome Sequence of Geobacillus stearothermophilus DSM 458, an Antimicrobial-Producing Thermophilic Bacterium, Isolated from a Sugar Beet Factory

2017 ◽  
Vol 5 (43) ◽  
Author(s):  
Kevin Egan ◽  
Philip Kelleher ◽  
Des Field ◽  
Mary C. Rea ◽  
R. Paul Ross ◽  
...  
Keyword(s):  
Sugar Beet ◽  
Genome Sequence ◽  
In Silico ◽  
Thermophilic Bacterium ◽  
Full Genome Sequence ◽  
Full Genome ◽  
Geobacillus Stearothermophilus ◽  
Biosynthetic Genes ◽  
Content Type

ABSTRACT This paper reports the full genome sequence of the antimicrobial-producing bacterium Geobacillus stearothermophilus DSM 458, isolated in a sugar beet factory in Austria. In silico analysis reveals the presence of a number of novel bacteriocin biosynthetic genes.

Novel environment friendly olive pomace-cyclohexanone formaldehyde composite resin

2021 ◽  
Vol ahead-of-print (ahead-of-print) ◽  
Author(s):  
Selda Sert ◽  
Nilgün Kızılcan
Keyword(s):  
Organic Solvents ◽  
Condensation Reaction ◽  
Agricultural Waste ◽  
Composite Resins ◽  
Formaldehyde Resin ◽  
Olive Pomace ◽  
Adhesion Properties ◽  
Content Type ◽  
Naoh Solution

Purpose Cyclohexanone-formaldehyde resin (CFR) was in situ modified with olive pomace (OP) in the presence of sodium hydroxide. The purpose of this study is to produce eco-friendly OP modified cyclohexanone composite resins (OPCFCR) with a one-step method that has higher condensation reaction temperature than CFR. The water absorption properties, gloss value and cross-cut adhesion properties of the product were investigated. Design/methodology/approach Cyclohexanone, formalin (37% aqueous solution) and tannin were mixed and 20% aqueous NaOH solution was added to produce the resin. OP has environmentally friendly bio-based lignin, cellulose and phenolic compounds and the OP structure has been incorporated into the structure of the CFR resin during the in situ modification, such as resole resin and polysaccharide. The weights of pomace were used as 5% and 10% of the weight of cyclohexanone in cyclohexanone-formaldehyde composite resins, respectively. Findings There is an improvement in the properties of the OPCFCR produced from an agricultural waste that is very abundant in Gulf of Edremit region of Balikesir. The OPCFCRs were soluble in common organic solvents. The product OPCFCR has a dark red-brown color. Research limitations/implications The reaction mixture must be stirred continuously. Subsequently, 37% formalin was added dropwise in total while refluxing. The amount of aqueous NaOH solution is limited as the formed resin may become insoluble in common organic solvents. At the end of the reaction, a water-insoluble resin is obtained. Practical implications This study provides the application of ketonic resins. The OPCFCR containing phenolic groups may also promote the adhesive strength of a coating. Social implications These resins may be used for the preparation of adhesive. OP, with a large amount of catechol groups, was considered for reducing the formaldehyde emission level on the adhesive system. Originality/value OPCFCR has been synthesized in the presence of a base catalyst. Environmental and ecological concerns have increased the attention paid by chemical industry to renewable raw materials.

Development of metaheuristics to solve a transportation inventory location routing problem considering lost sale for perishable goods

2019 ◽  
Vol 14 (1) ◽  
pp. 175-198 ◽  
Author(s):  
S.M.T. Fatemi Ghomi ◽  
B. Asgarian
Keyword(s):  
Mathematical Model ◽  
Distribution Systems ◽  
Hybrid Approach ◽  
Rational Approach ◽  
Location Routing Problem ◽  
Routing Problem ◽  
Content Type ◽  
Location Routing ◽  
Perishable Goods ◽  
The Impact

PurposeFinding a rational approach to maintain a freshness of foods and perishable goods and saving their intrinsic attributes during a distribution of these products is one of the main issues for distribution and logistics companies. This paper aims to provide a framework for distribution of perishable goods which can be applied for real life situations.Design/methodology/approachThis paper proposes a novel mathematical model for transportation inventory location routing problem. In addition, the paper addresses the impact of perishable goods age on the demand of final customers. The model is optimally solved for small- and medium-scale problems. Moreover, regarding to NP-hard nature of the proposed model, two simple and one hybrid metaheuristic algorithms are developed to cope with the complexity of problem in large scale problems.FindingsNumerical examples with different scenarios and sensitivity analysis are conducted to investigate the performance of proposed algorithms and impacts of important parameters on optimal solutions. The results show the acceptable performance of proposed algorithms.Originality/valueThe authors formulate a novel mathematical model which can be applicable in perishable goods distribution systems In this regard, the authors consider lost sale which is proportional to age of products. A new hybrid approach is applied to tackle the problem and the results show the rational performance of the algorithm.

scholarly journals The interplay between SME owner-managers and the brand-as-a-person

2019 ◽  
Vol 28 (4) ◽  
pp. 555-572 ◽  
Author(s):  
Edgar Centeno ◽  
Jesus Cambra-Fierro ◽  
Rosario Vazquez-Carrasco ◽  
Susan J. Hart ◽  
Keith Dinnie
Keyword(s):  
Personality Traits ◽  
Learning Strategy ◽  
Personal Characteristics ◽  
Rational Approach ◽  
Theory Approach ◽  
Structured Interviews ◽  
Strategy Formulation ◽  
Content Type ◽  
Brand Strategies ◽  
Grounded Theory Approach

PurposeThe purpose of this paper is to investigate the largely unexplored conceptualisation of the brand-as-a-person metaphor in small-to-medium-sized enterprises (SMEs) by examining its potential relation with the SME owner-manager, the pathways to its creation and development and the intuitive nature of this relationship.Design/methodology/approachA grounded theory approach was used, and data were collected through a set of 36 semi-structured interviews with 30 SME owner-managers in various sectors in Mexico.FindingsThe results indicate that SME owner-managers intuitively humanise their brands. The study revealed four pathways to develop the brand-as-a-person metaphor in the SME context: through personality traits, tastes and preferences, abilities and knowledge and values, all suggesting that SMEs’ brand-as-a-person metaphors are largely an extension of their owner-managers.Research limitations/implicationsThe paper presents a theoretical framework that illustrates the four pathways to the creation and development of brand-as-a-person that are derived from the brand’s relationship with the SME owner-manager. The results of cross-industry semi-structured interviews are limited to a single culture context.Practical implicationsSME owner-managers should first undertake an introspective personal assessment of their intuitive and conscious decision-making, as SME owner-managers often make decisions in an intuitive way. The results suggest that they should act in a more conscious, responsible and rational way when formulating their brand strategies.Originality/valueThis is the first study to clarify the profound influence of SME owner-managers’ personal characteristics, including personality traits, tastes and preferences, abilities and knowledge and values, on the brand-as-a-person metaphor. This study also confirms the intuitive learning strategy formulation of SME owner-managers’ branding practices and SMEs’ need for a more rational approach to branding.

Factors affecting Vietnamese farmers' intention toward organic agricultural production

2021 ◽  
Vol ahead-of-print (ahead-of-print) ◽  
Author(s):  
Thi Phuong Linh Nguyen ◽  
Xuan Hau Doan ◽  
Thu Thuy Nguyen ◽  
Thi Mai Nguyen
Keyword(s):  
Factor Analysis ◽  
Organic Farming ◽  
Organic Agriculture ◽  
Agricultural Production ◽  
Subjective Norms ◽  
Rational Approach ◽  
Research Model ◽  
Content Type ◽  
Factors Affecting ◽  
Personal Norm

PurposeThe purpose of this paper is to investigate factors affecting Vietnamese farmer's intention toward organic agricultural production based on research model integrating theories: theory of planned behavior (TPB) and norm activation model (NAM).Design/methodology/approachAfter in-depth interviews with 5 agricultural researchers and 5 farmers, the authors determined the official research model and built a complete survey. Data were collected from 318 farmers in the Hanoi, Vietnam by directly survey. Statistical methods, such as Cronbach's alpha analysis, exploratory factor analysis (EFA), confirmatory factor analysis (CFA) and structural equation modeling (SEM), were used to analyze the data.FindingsThis paper shows that attitude, subjective norms, perceived behavioral control and personal norm were significantly related to intention of the farmers; awareness of consequences was positively related to ascription of responsibility, personal norm, attitude and subjective norms; ascription of responsibility has a positive relationship with personal norm. The TPB-NAM integration model is proven to be superior to the original TPB model when studying factors affecting Vietnamese farmer's intention toward organic agricultural production.Research limitations/implicationsThe main limitation of this paper is that the sampling method is not representative for the whole country and just stops at researching the intentions of farmers without understanding the practices of organic agricultural production.Practical implicationsThe findings indicate that state management agencies in Vietnam need to pay attention to raise awareness among farmers about the role of organic farming, communicate to farmers through different channels about the benefits of organic farming compared to conventional agriculture and form information spillover groups between farmers who have produced organic agriculture and have not yet produced organic agriculture.Originality/valueThis paper contributes to the existing literature by focusing on integrating TPB and NAM when understanding farmers' intention toward organic agricultural production in Vietnam. This integrated model has proven the suitability of combining two approaches, a rational approach and an ethical approach, when studying farmer intentions.

scholarly journals Chromosome Engineering To Generate Plasmid-Free Phenylalanine- and Tyrosine-Overproducing Escherichia coli Strains That Can Be Applied in the Generation of Aromatic-Compound-Producing Bacteria

2020 ◽  
Vol 86 (14) ◽  
Author(s):  
Daisuke Koma ◽  
Takahiro Kishida ◽  
Eisuke Yoshida ◽  
Hiroyuki Ohashi ◽  
Hayato Yamanaka ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Aromatic Compound ◽  
Aromatic Compounds ◽  
High Yield ◽  
Rational Approach ◽  
Chromosome Engineering ◽  
Phenyllactic Acid ◽  
Content Type ◽  
E Coli ◽  
Free Strains

ABSTRACT Many phenylalanine- and tyrosine-producing strains have used plasmid-based overexpression of pathway genes. The resulting strains achieved high titers and yields of phenylalanine and tyrosine. Chromosomally engineered, plasmid-free producers have shown lower titers and yields than plasmid-based strains, but the former are advantageous in terms of cultivation cost and public health/environmental risk. Therefore, we engineered here the Escherichia coli chromosome to create superior phenylalanine- and tyrosine-overproducing strains that did not depend on plasmid-based expression. Integration into the E. coli chromosome of two central metabolic pathway genes (ppsA and tktA) and eight shikimate pathway genes (aroA, aroB, aroC, aroD, aroE, aroGfbr, aroL, and pheAfbr), controlled by the T7lac promoter, resulted in excellent titers and yields of phenylalanine; the superscript “fbr” indicates that the enzyme encoded by the gene was feedback resistant. The generated strain could be changed to be a superior tyrosine-producing strain by replacing pheAfbr with tyrAfbr. A rational approach revealed that integration of seven genes (ppsA, tktA, aroA, aroB, aroC, aroGfbr, and pheAfbr) was necessary as the minimum gene set for high-yield phenylalanine production in E. coli MG1655 (tyrR, adhE, ldhA, pykF, pflDC, and ascF deletant). The phenylalanine- and tyrosine-producing strains were further applied to generate phenyllactic acid-, 4-hydroxyphenyllactic acid-, tyramine-, and tyrosol-producing strains; yield of these aromatic compounds increased proportionally to the increase in phenylalanine and tyrosine yields. IMPORTANCE Plasmid-free strains for aromatic compound production are desired in the aspect of industrial application. However, the yields of phenylalanine and tyrosine have been considerably lower in plasmid-free strains than in plasmid-based strains. The significance of this research is that we succeeded in generating superior plasmid-free phenylalanine- and tyrosine-producing strains by engineering the E. coli chromosome, which was comparable to that in plasmid-based strains. The generated strains have a potential to generate superior strains for the production of aromatic compounds. Actually, we demonstrated that four kinds of aromatic compounds could be produced from glucose with high yields (e.g., 0.28 g tyrosol/g glucose).

A rational approach to identify and cluster intangible assets

2015 ◽  
Vol 16 (4) ◽  
pp. 809-834 ◽  
Author(s):  
Franco M. Battagello ◽  
Michele Grimaldi ◽  
Livio Cricelli
Keyword(s):  
Business Performance ◽  
Intangible Assets ◽  
Mutual Influence ◽  
Building Blocks ◽  
Rational Approach ◽  
Content Type ◽  
Knowledge Based ◽  
Strategic Resources ◽  
Strategy Alignment ◽  
A Company

Purpose – This study is intended to work out a bottleneck in the comprehension of the relational nexus which links the set of key strategic resources (SRs) of a company, represented by the uncertain recognition and the ambiguous clustering of their intangible components. The purpose of this paper is to provide a candidate solution for a rational appraisal of the inventory of the knowledge-based resources held by a company, which synergically form its Intellectual Capital (IC). Design/methodology/approach – This goal is achieved by the means of a qualitative/quantitative approach composed of sequential phases, intended to: atomize the value domain of the firm into its basic building blocks; gauge their mutual interactions and impacts; re-aggregate those involved entities accordingly; cluster them into a collection of identified and validated Intangible Assets (IAs). Never giving any direct judgment on the IAs themselves (whose extension can be fuzzy or unknown). But on the impacts between the value drivers they are built on. Findings – The proposed procedure, step-by-step illustrated by means of a numerical simulation, out of the amorphous mass of the SRs, returns an analytic picture of its composing elements keeping track of their intertwined connections and mutual influence. Consequently, allowing the comprehension of the actual framing and of the relational positioning and magnitude of such entities. Practical implications – This risk-mitigated rational identification of IAs allows the analyst to target a proper evaluation technique on them. And the management of the company to mindfully allocate/leverage on them to improve business performance and strategy alignment. The implementation returns some analytic tools which render a diagnostic snapshot of the composing elements of the IC, increasing the awareness of such entities and allowing internal/external benchmarking. Originality/value – The suggested methodology mitigates the risk of discretionality in the definition of the perimeter of each target-entity, by avoiding any direct biased judgment on them. So that each asset gets unambiguously identified within a network-logic and the interlinked portfolio of knowledge-based resources can be assessed and managed in an rational and traceable way.

scholarly journals Explaining the premium to NAV in publicly traded Australian REITs, 2008–2018

2019 ◽  
Vol 38 (1) ◽  
pp. 4-30 ◽  
Author(s):  
Isil Erol ◽  
Tanja Tyvimaa
Keyword(s):  
Financial Crisis ◽  
Real Estate ◽  
Global Financial Crisis ◽  
Rational Approach ◽  
Annual Data ◽  
Financial Leverage ◽  
Publicly Traded ◽  
Aggressive Approach ◽  
Content Type ◽  
Equity Ratio

Purpose The purpose of this paper is to explore the levels and determinants of net asset value (NAV) premiums/discounts for publicly traded Australian Real Estate Investment Trust (A-REIT) market during the last decade. A-REITs were severely affected by the global financial crisis as S&P/ASX 200 A-REIT index-listed property stocks experienced 47 per cent discount to NAV, on average, in 2008–2009 crisis. Since 2013, A-REIT sector has exhibited a strong recovery from the financial crisis and traded at high premiums to date. Understanding the relationship between pricing in the public and private real estate markets has taken on great importance as A-REITs continue to trade at significant premium to NAV unlike their counterparts in the USA and Europe. Design/methodology/approach This paper follows a rational approach to explain variations in NAV premiums and explores the company-specific factors such as liquidity, financial leverage, size, stock price volatility and portfolio diversification behind the A-REIT NAV premiums/discounts. The study specifies and estimates a model of cross-sectional and time variation in premiums/discounts to NAV using semi-annual data for a sample of 40 A-REITs over the 2008–2018 period. Findings The results reveal that A-REIT premiums to NAV can be explained not only by the liquidity benefit of listed property stocks but also positive financial leverage effect. During the past decade, A-REITs have followed an aggressive approach in financing their growth by using borrowed funds to purchase assets as the income from the property offsets the cost of borrowing and the risk that accompanies it. Debt-to-equity ratio has to be considered as an important source of NAV premiums as highly geared A-REITs that favoured debt financing over equity financing traded at significant premiums to NAV of their underlying real estate assets. Practical implications The paper includes implications for the REIT market investors. The regression analysis shows that specialty A-REITs with a focus on creative market niches traded at higher premiums compared with other property stocks, especially in the post-GFC recovery period. Specialty REITs are more highly valued by the market than their traditional specialised counterparts (e.g. office and retail REITs), and those pursuing a diversified strategy. Originality/value This paper presents an Australian case study as the A-REIT market provides a suitable environment for testing the effect of financial gearing on the REIT premium to NAV. The study provides empirical evidence supporting the importance of debt-to-equity ratio in explaining the variation in A-REIT NAV premiums.

scholarly journals Whole-Genome Sequences of Five Geobacillus stearothermophilus Strains Isolated from Processing Lines of Powdered Infant Formula

2019 ◽  
Vol 8 (4) ◽  
Author(s):  
Tae Jin Cho ◽  
Dongmin Yang ◽  
Byeonghyeok Park ◽  
In-Geol Choi ◽  
Min Suk Rhee
Keyword(s):  
Infant Formula ◽  
Safety Management ◽  
Whole Genome ◽  
Powdered Infant Formula ◽  
Geobacillus Stearothermophilus ◽  
Genome Sequences ◽  
Content Type ◽  
Manufacturing Plants ◽  
Work In Process ◽  
Concentrated Milk

Geobacillus stearothermophilus is the thermophile present in processing lines of powdered infant formula (PIF). We report the whole-genome sequences of G. stearothermophilus strains isolated from work-in-process products (sterilized and concentrated milk) of manufacturing plants. Understanding the genomic basis governing the metabolism of G. stearothermophilus can contribute to the safety management of PIF during its manufacture.

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