Escherichia coli and Pichia pastoris: microbial cell-factory platform for -full-length IgG production.

Owing to the unmet demand, the pharmaceutical industry is investigating an alternative host to mammalian cells to produce antibodies for a variety of therapeutic and research applications. Regardless of some disadvantages, Escherichia coli and Pichia pastoris are the preferred microbial hosts for antibody production. Despite the fact that the production of full-length antibodies has been successfully demonstrated in E. coli, which has mostly been used to produce antibody fragments, such as: antigen-binding fragments (Fab), single-chain fragment variable (scFv), and nanobodies. In contrast, Pichia, a eukaryotic microbial host, is mostly used to produce glycosylated full-length antibodies, though hypermannosylated glycan is a major challenge. Advanced strategies, such as the introduction of human-like glycosylation in endotoxin-edited E. coli and cell-free system-based glycosylation, are making progress in creating human-like glycosylation profiles of antibodies in these microbes. This review begins by explaining the structural and functional requirements of antibodies and continues by describing and analyzing the potential of E. coli and P. pastoris as hosts for providing a favorable environment to create a fully functional antibody. In addition, authors compare these microbes on certain features and predict their future in antibody production. Briefly, this review analyzes, compares, and highlights E. coli and P. pastoris as potential hosts for antibody production.

Functional cooperation of the glycine synthase-reductase and Wood-Ljungdahl pathways for autotrophic growth of Clostridium drakei.

Among CO2-fixing metabolic pathways in nature, the linear Wood-Ljungdahl pathway (WLP) in phylogenetically diverse acetate-forming acetogens comprises the most energetically efficient pathway, requires the least number of reactions, and converts CO2 to formate and then into acetyl-CoA. Despite two genes encoding glycine synthase being well-conserved in WLP gene clusters, the functional role of glycine synthase under autotrophic growth conditions has remained uncertain. Here, using the reconstructed genome-scale metabolic model iSL771 based on the completed genome sequence, transcriptomics, 13C isotope-based metabolite-tracing experiments, biochemical assays, and heterologous expression of the pathway in another acetogen, we discovered that the WLP and the glycine synthase pathway are functionally interconnected to fix CO2, subsequently converting CO2 into acetyl-CoA, acetyl-phosphate, and serine. Moreover, the functional cooperation of the pathways enhances CO2 consumption and cellular growth rates via bypassing reducing power required reactions for cellular metabolism during autotrophic growth of acetogens.

Development of Methylorubrum extorquens AM1 as a promising platform strain for enhanced violacein production from co-utilization of methanol and acetate.

Violacein, a blue-violet compound with a wide range of beneficial bioactivities, is an attractive product for microbial production. Currently, violacein production has been demonstrated in several sugar heterotrophs through metabolic engineering; however, the cost of production remains an obstacle for business ventures. To address this issue, the development of host strains that can utilize inexpensive alternative substrates to reduce production costs would enable the commercialization of violacein. In this study, we engineered a facultative methylotroph, Methylorubrum extorquens AM1, to develop a methanol-based platform for violacein production. By optimizing expression vectors as well as inducer concentrations, 11.7 mg/L violacein production was first demonstrated using methanol as the sole substrate. Considering that unidentified bottlenecks for violacein biosynthesis in the shikimate pathway of M. extorquens AM1 would be difficult to address using generic metabolic engineering approaches, random mutagenesis and site-directed mutagenesis were implemented, and a 2-fold improvement in violacein production was achieved. Finally, by co-utilization of methanol and acetate, a remarkable enhancement of violacein production to 118 mg/L was achieved. Our results establish a platform strain for violacein production from non-sugar feedstocks, which may contribute to the development of an economically efficient large-scale fermentation system for violacein production.

Unlocking the biosynthesis of sesquiterpenoids from methane via the methylerythritol phosphate pathway in methanotrophic bacteria, using α-humulene as a model compound.

Isoprenoids are an abundant and diverse class of natural products with various applications in the pharmaceutical, cosmetics and biofuel industries. A methanotroph-based biorefinery is an attractive scenario for the production of a variety of value-added compounds from methane, because methane is a promising alternative feedstock for industrial biomanufacturing. In this study, we metabolically engineered Methylotuvimicrobium alcaliphilum 20Z for de novo synthesis of a sesquiterpenoid from methane, using α-humulene as a model compound, via optimization of the native methylerythritol phosphate (MEP) pathway. Expression of codon-optimized α-humulene synthase from Zingiber zerumbet in M. alcaliphilum 20Z resulted in an initial yield of 0.04 mg/g dry cell weight. Overexpressing key enzymes (IspA, IspG, and Dxs) for debottlenecking of the MEP pathway increased α-humulene production 5.2-fold compared with the initial strain. Subsequently, redirecting the carbon flux through the Embden-Meyerhof-Parnas pathway resulted in an additional 3-fold increase in α-humulene production. Additionally, a genome-scale model using flux scanning based on enforced objective flux method was used to identify potential overexpression targets to increase flux towards isoprenoid production. Several target reactions from cofactor synthesis pathways were probed and evaluated for their effects on α-humulene synthesis, resulting in α-humulene yield up to 0.75 mg/g DCW with 18.8-fold enhancement from initial yield. This study first demonstrates production of a sesquiterpenoid from methane using methanotrophs as the biocatalyst and proposes potential strategies to enhance production of sesquiterpenoid and related isoprenoid products in engineered methanotrophic bacteria.

Metabolic engineering of type II methanotroph, Methylosinus trichosporium OB3b, for production of 3-hydroxypropionic acid from methane via a malonyl-CoA reductase-dependent pathway.

We engineered a type II methanotroph, Methylosinus trichosporium OB3b, for 3-hydroxypropionic acid (3HP) production by reconstructing malonyl-CoA pathway through heterologous expression of Chloroflexus aurantiacus malonyl-CoA reductase (MCR), a bifunctional enzyme. Two strategies were designed and implemented to increase the malonyl-CoA pool and thus, increase in 3HP production. First, we engineered the supply of malonyl-CoA precursors by overexpressing endogenous acetyl-CoA carboxylase (ACC), substantially enhancing the production of 3HP. Overexpression of biotin protein ligase (BPL) and malic enzyme (NADP+-ME) led to a ∼22.7% and ∼34.5% increase, respectively, in 3HP titer in ACC-overexpressing cells. Also, the acetyl-CoA carboxylation bypass route was reconstructed to improve 3HP productivity. Co-expression of methylmalonyl-CoA carboxyltransferase (MMC) of Propionibacterium freudenreichii and phosphoenolpyruvate carboxylase (PEPC), which provides the MMC precursor, further improved the 3HP titer. The highest 3HP production of 49 mg/L in the OB3b-MCRMP strain overexpressing MCR, MMC and PEPC resulted in a 2.4-fold improvement of titer compared with that in the only MCR-overexpressing strain. Finally, we could obtain 60.59 mg/L of 3HP in 42 h using the OB3b-MCRMP strain through bioreactor operation, with a 6.36-fold increase of volumetric productivity compared than that in the flask cultures. This work demonstrates metabolic engineering of type II methanotrophs, opening the door for using type II methanotrophs as cell factories for biochemical production along with mitigation of greenhouse gases.

Genome-scale evaluation of core one-carbon metabolism in gammaproteobacterial methanotrophs grown on methane and methanol.

Methylotuvimicrobium alcaliphilum 20Z is a promising platform strain for bioconversion of one-carbon (C1) substrates into value-added products. To carry out robust metabolic engineering with methylotrophic bacteria and to implement C1 conversion machinery in non-native hosts, systems-level evaluation and understanding of central C1 metabolism in methanotrophs under various conditions is pivotal but yet elusive. In this study, a genome-scale integrated approach was used to provide in-depth knowledge on the metabolic pathways of M. alcaliphilum 20Z grown on methane and methanol. Systems assessment of core carbon metabolism indicated the methanol assimilation pathway is mostly coupled with the efficient Embden-Meyerhof-Parnas (EMP) pathway along with the serine cycle. In addition, an incomplete TCA cycle operated in M. alcaliphilum 20Z on methanol, which might only supply precursors for de novo synthesis but not reducing powers. Instead, it appears that the direct formaldehyde oxidation pathway supply energy for the whole metabolic system. Additionally, a comparative transcriptomic analysis in multiple gammaproteobacterial methanotrophs also revealed the transcriptional responses of central metabolism on carbon substrate change. These findings provided a systems-level understanding of carbon metabolism and new opportunities for strain design to produce relevant products from different C1-feedstocks.

Metabolic role of pyrophosphate-linked phosphofructokinase pfk for C1 assimilation in Methylotuvimicrobium alcaliphilum 20Z.

BACKGROUND: Methanotrophs is a promising biocatalyst in biotechnological applications with their ability to utilize single carbon (C1) feedstock to produce high-value compounds. Understanding the behavior of biological networks of methanotrophic bacteria in different parameters is vital to systems biology and metabolic engineering. Interestingly, methanotrophic bacteria possess the pyrophosphate-dependent 6-phosphofructokinase (PPi-PFK) instead of the ATP-dependent 6-phosphofructokinase, indicating their potentials to serve as promising model for investigation the role of inorganic pyrophosphate (PPi) and PPi-dependent glycolysis in bacteria. Gene knockout experiments along with global-omics approaches can be used for studying gene functions as well as unraveling regulatory networks that rely on the gene product. RESULTS: In this study, we performed gene knockout and RNA-seq experiments in Methylotuvimicrobium alcaliphilum 20Z to investigate the functional roles of PPi-PFK in C1 metabolism when cells were grown on methane and methanol, highlighting its metabolic importance in C1 assimilation in M. alcaliphilum 20Z. We further conducted adaptive laboratory evolution (ALE) to investigate regulatory architecture in pfk knockout strain. Whole-genome resequencing and RNA-seq approaches were performed to characterize the genetic and metabolic responses of adaptation to pfk knockout. A number of mutations, as well as gene expression profiles, were identified in pfk ALE strain to overcome insufficient C1 assimilation pathway which limits the growth in the unevolved strain. CONCLUSIONS: This study first revealed the regulatory roles of PPi-PFK on C1 metabolism and then provided novel insights into mechanism of adaptation to the loss of this major metabolic enzyme as well as an improved basis for future strain design in type I methanotrophs.

Developments of Riboswitches and Toehold Switches for Molecular Detection-Biosensing and Molecular Diagnostics.

Riboswitches and toehold switches are considered to have potential for implementation in various fields, i.e., biosensing, metabolic engineering, and molecular diagnostics. The specific binding, programmability, and manipulability of these RNA-based molecules enable their intensive deployments in molecular detection as biosensors for regulating gene expressions, tracking metabolites, or detecting RNA sequences of pathogenic microorganisms. In this review, we will focus on the development of riboswitches and toehold switches in biosensing and molecular diagnostics. This review introduces the operating principles and the notable design features of riboswitches as well as toehold switches. Moreover, we will describe the advances and future directions of riboswitches and toehold switches in biosensing and molecular diagnostics.

A comparative transcriptome analysis of the novel obligate methanotroph Methylomonas sp. DH-1 reveals key differences in transcriptional responses in C1 and secondary metabolite pathways during growth on methane and methanol.

BACKGROUND: Methanotrophs play an important role in biotechnological applications, with their ability to utilize single carbon (C1) feedstock such as methane and methanol to produce a range of high-value compounds. A newly isolated obligate methanotroph strain, Methylomonas sp. DH-1, became a platform strain for biotechnological applications because it has proven capable of producing chemicals, fuels, and secondary metabolites from methane and methanol. In this study, transcriptome analysis with RNA-seq was used to investigate the transcriptional change of Methylomonas sp. DH-1 on methane and methanol. This was done to improve knowledge about C1 assimilation and secondary metabolite pathways in this promising, but under-characterized, methane-bioconversion strain. RESULTS: We integrated genomic and transcriptomic analysis of the newly isolated Methylomonas sp. DH-1 grown on methane and methanol. Detailed transcriptomic analysis indicated that (i) Methylomonas sp. DH-1 possesses the ribulose monophosphate (RuMP) cycle and the Embden-Meyerhof-Parnas (EMP) pathway, which can serve as main pathways for C1 assimilation, (ii) the existence and the expression of a complete serine cycle and a complete tricarboxylic acid (TCA) cycle might contribute to methane conversion and energy production, and (iii) the highly active endogenous plasmid pDH1 may code for essential metabolic processes. Comparative transcriptomic analysis on methane and methanol as a sole carbon source revealed different transcriptional responses of Methylomonas sp. DH-1, especially in C1 assimilation, secondary metabolite pathways, and oxidative stress. Especially, these results suggest a shift of central metabolism when substrate changed from methane to methanol in which formaldehyde oxidation pathway and serine cycle carried more flux to produce acetyl-coA and NADH. Meanwhile, downregulation of TCA cycle when grown on methanol may suggest a shift of its main function is to provide de novo biosynthesis, but not produce NADH. CONCLUSIONS: This study provides insights into the transcriptomic profile of Methylomonas sp. DH-1 grown on major carbon sources for C1 assimilation, providing in-depth knowledge on the metabolic pathways of this strain. These observations and analyses can contribute to future metabolic engineering with the newly isolated, yet under-characterized, Methylomonas sp. DH-1 to enhance its biochemical application in relevant industries.

Metabolic engineering of the type I methanotroph Methylomonas sp. DH-1 for production of succinate from methane.

Methane-utilizing methanotrophs are fascinating systems for methane bioconversion. Methylomonas sp. DH-1, a novel type I methanotroph isolated from brewery sludge, has been evaluated as a promising candidate for an industrial bio-catalyst. Succinate has been considered one of the top building block chemicals for the agricultural, food, and pharmaceutical industries. In this study, Methylomonas sp. DH-1 was engineered to accumulate succinate as a desired product. The TCA cycle and enzymes diverting carbon flux to acetate or formate were modified or deleted to improve succinate productivity. By deleting succinate dehydrogenase (sdh) in the TCA cycle, succinate production increased dramatically ∼10 times compared to that of the wild type. In addition, the maximum succinate titer of ∼134 mg/L (DS-GL) was achieved by integrating glyoxylate shunt enzymes from the E. coli MG1655 strain. Pyruvate formate lyase (pfl) and acetate kinase-phosphotransacetylase (ack-pta) genes were disrupted to further concentrate carbon flux to the TCA cycle. However, these additional disruptions of competitive pathways did not affect cell growth or succinate production positively. The mutant strain DS-GL, which showed the best succinate production, was grown in a fed-batch bioreactor, and higher cell growth and succinate production (∼195 mg/L succinate with 0.0789 g-succinate/g-methane yield) were achieved. In this study, we demonstrated a novel platform for microbial conversion of methane to succinate using methanotroph.

Biological conversion of propane to 2-propanol using group I and II methanotrophs as biocatalysts.

Propane is the main component of liquefied petroleum gas and is derived from crude oil processing. Methanotrophic bacteria can convert various alkanes using methane monooxygenase enzyme to primary alcohols. These are further oxidized to various aldehydes by alcohol dehydrogenases or methanol dehydrogenases. In this study, 2-propanol was produced from propane using the whole cells of Methylosinus trichosporium OB3b, Methylomicrobium alcaliphilum 20Z, and Methylomonas sp. DH-1 as the biocatalysts. The biocatalytic process of converting propane to 2-propanol was optimized by the use of several inhibitors and additives, such as EDTA, sodium phosphate, and sodium formate to prevent oxidation of 2-propanol to acetone and to enhance conversion of propane to propanol. The maximum titer of 2-propanol was 0.424 g/L, 0.311 g/L, and 0.610 g/L for Methylomonas sp. DH-1, M. alcaliphilum 20Z, and M. trichosporium OB3b whole cells, respectively. These results showed that type I and type II methanotrophs could be used as the potent biocatalyst for conversion of propane to propanol.

Development and Optimization of the Biological Conversion of Ethane to Ethanol Using Whole-Cell Methanotrophs Possessing Methane Monooxygenase.

The biological production of ethanol from ethane for the utilization of ethane in natural gas was investigated under ambient conditions using whole-cell methanotrophs possessing methane monooxygenase. Several independent variables including ethane concentration and biocatalyst amounts, among other factors, were optimized for the enhancement of ethane-to-ethanol bioconversion. We obtained 0.4 g/L/h of volumetric productivity and 0.52 g/L of maximum titer in optimum batch reaction conditions. In this study, we demonstrate that the biological gas-to-liquid conversion of ethane to ethanol has potent technical feasibility as a new application of ethane gas.

Systematic metabolic engineering of Methylomicrobium alcaliphilum 20Z for 2,3-butanediol production from methane.

Methane is considered a next-generation feedstock, and methanotrophic cell-based biorefinery is attractive for production of a variety of high-value compounds from methane. In this work, we have metabolically engineered Methylomicrobium alcaliphilum 20Z for 2,3-butanediol (2,3-BDO) production from methane. The engineered strain 20Z/pBudK.p, harboring the 2,3-BDO synthesis gene cluster (budABC) from Klebsiella pneumoniae, accumulated 2,3-BDO in methane-fed shake flask cultures with a titer of 35.66 mg/L. Expression of the most efficient gene cluster was optimized using selection of promoters, translation initiation rates (TIR), and the combination of 2,3-BDO synthesis genes from different sources. A higher 2,3-BDO titer of 57.7 mg/L was measured in the 20Z/pNBM-Re strain with budA of K. pneumoniae and budB of Bacillus subtilis under the control of the Tac promoter. The genome-scale metabolic network reconstruction of M. alcaliphilum 20Z enabled in silico gene knockout predictions using an evolutionary programming method to couple growth and 2,3-BDO production. The ldh, ack, and mdh genes in M. alcaliphilum 20Z were identified as potential knockout targets. Pursuing these targets, a triple-mutant strain ∆ldh ∆ack ∆mdh was constructed, resulting in a further increase of the 2,3-BDO titer to 68.8 mg/L. The productivity of this optimized strain was then tested in a fed-batch stirred tank bioreactor, where final product concentrations of up to 86.2 mg/L with a yield of 0.0318 g-(2,3-BDO) /g-CH4 were obtained under O2-limited conditions. This study first demonstrates the strategy of in silico simulation-guided metabolic engineering and represents a proof-of-concept for the production of value-added compounds using systematic approaches from engineered methanotrophs.

Biological conversion of methane to chemicals and fuels: technical challenges and issues.

Methane is a promising next-generation carbon feedstock for industrial biotechnology due to its low price and huge availability. Biological conversion of methane to valuable products can mitigate methane-induced global warming as greenhouse gas. There have been challenges for the conversion of methane into various chemicals and fuels using engineered non-native hosts with synthetic methanotrophy or methanotrophs with the reconstruction of synthetic pathways for target products. Herein, we analyze the technical challenges and issues of potent methane bioconversion technology. Pros and cons of metabolic engineering of methanotrophs for methane bioconversion, and perspectives on the bioconversion of methane to chemicals and liquid fuels are discussed.

Growth of Silver Nanowires from Controlled Silver Chloride Seeds and Their Application for Fluorescence Enhancement Based on Localized Surface Plasmon Resonance.

A "Polyol" method has granted low-cost and facile process-controllability for silver-nanowire (Ag-NW) synthesis. Although homogenous and heterogeneous nucleation and growth during Ag-NW synthesis are possible using polyol methods, heterogeneous nucleation and growth of Ag NW guarantees highly selective growth of nanostructures using silver chloride (AgCl) seeds, which provides a stable source of chloride ions (Cl-) and thermodynamic reversibility. In this paper, a microdroplet has been adopted to synthesize uniform AgCl seeds with different diameter that are used for seed-mediated Ag-NW synthesis. The concentration of two precursors (AgNO3 and NaCl) in the droplets is modulated to produce different sizes of AgCl seeds, which determines the diameter and length of Ag NWs. The process of the seed-mediated growth of Ag NWs has been monitored by observing the peak shift in the time-resolved UV-vis extinction spectrum. Furthermore, the distinct plasmonic property of Ag NWs for transverse and longitudinal localized-surface-plasmon-resonance (LSPR)-mediated fluorescence enhancement is utilized. The high aspect ratio and sharp tips work as simple antennas that induce the enhanced fluorescence emission intensity of a fluorophore, which can be applied in the fields of biological tissue imaging and therapy.

Selective bio-oxidation of propane to acetone using methane-oxidizing Methylomonas sp. DH-1.

Propane is the major component of liquefied petroleum gas (LPG). Nowadays, the use of LPG is decreasing, and thus utilization of propane as a chemical feedstock is in need of development. An efficient biological conversion of propane to acetone using a methanotrophic whole cell as the biocatalyst was proposed and investigated. A bio-oxidation pathway of propane to acetone in Methylomonas sp. DH-1 was analyzed by gene expression profiling via RNA sequencing. Propane was oxidized to 2-propanol by particulate methane monooxygenase and subsequently to acetone by methanol dehydrogenases. Methylomonas sp. DH-1 was deficient in acetone-converting enzymes and thus accumulated acetone in the absence of any enzyme inhibition. The maximum accumulation, average productivity and specific productivity of acetone were 16.62 mM, 0.678 mM/h and 0.141 mmol/g cell/h, respectively, under the optimized conditions. Our study demonstrates a novel method for the bioconversion of propane to acetone using methanotrophs under mild reaction condition.

Characterization of the Two Methylation Steps Involved in the Biosynthesis of Mycinose in Tylosin.

The S-adenosyl-l-methionine-dependent O-methyltransferases TylE and TylF catalyze the last two methylation reactions in the tylosin biosynthetic pathway of Streptomyces fradiae. It has long been known that the TylE-catalyzed C2‴-O-methylation of the 6-deoxy-d-allose bound to demethylmacrocin or demethyllactenocin precedes the TylF-catalyzed C3‴-O-methylation of the d-javose (C2‴-O-methylated 6-deoxy-d-allose) attached to macrocin or lactenocin. This study reveals the unexpected substrate promiscuity of TylE and TylF responsible for the biosynthesis of d-mycinose (C3‴-O-methylated d-javose) in tylosin through the identification of a new minor intermediate 2‴-O-demethyldesmycosin (2; 3‴-methyl-demethyllactenocin), which lacks a 2‴-O-methyl group on the mycinose moiety of desmycosin, along with 2‴-O-demethyltylosin (1; 3‴-methyl-demethylmacrocin) that was previously detected from the S. fradiae mutant containing a mutation in the tylE gene. These results unveil the unique substrate flexibility of TylE and TylF and demonstrate their potential for the engineered biosynthesis of novel glycosylated macrolide derivatives.

Stabilization and fabrication of microbubbles: applications for medical purposes and functional materials.

Microbubbles with diameters ranging from a few micrometers to tens of micrometers have garnered significant attention in various applications including food processing, water treatment, enhanced oil recovery, surface cleaning, medical purposes, and material preparation fields with versatile functionalities. A variety of techniques have been developed to prepare microbubbles, such as ultrasonication, excimer laser ablation, high shear emulsification, membrane emulsification, an inkjet printing method, electrohydrodynamic atomization, template layer-by-layer deposition, and microfluidics. Generated bubbles should be immediately stabilized via the adsorption of stabilizing materials (e.g., surfactants, lipids, proteins, and solid particles) onto the gas-liquid interface to lower the interfacial tension. Such adsorption of stabilizers prevents coalescence between the microbubbles and also suppresses gas dissolution and resulting disproportionation caused by the presence of the Laplace overpressure across the gas-liquid interface. Herein, we comprehensively review three important topics of microbubbles: stabilization, fabrication, and applications.

Molecular cloning and characterization of a novel acetylalginate esterase gene in alg operon from Sphingomonas sp. MJ-3.

A novel acetylalginate esterase (AcAlgE) gene was cloned and characterized from the genomic DNA library of Sphingomonas sp. MJ-3. A putative gene encoding AcAlgE protein of 292-residue precursor protein with 20-amino acid signal peptide was identified in the alg operon. The deduced AcAlgE protein has GDSL-like consensus motif and shares a highest sequence identity (51%) with GDSL family lipolytic protein from Pseudoxanthomonas suwonensis. Enzymatic assays with bacterial acetylalginate as the substrate showed that the recombinant AcAlgE protein possesses deacetylation activity. The optimal temperature and pH for the AcAlgE were 22 °C and pH 6.5 (citrate buffer), respectively. The recombinant AcAlgE protein catalyzed deacetylation of acetylalginate with release of acetate. The resulting de-acetylated alginate was readily degraded by alginate lyases, indicating that the recombinant AcAlgE enhanced the subsequent degradation of acetylalginate by alginate lyases. The recombinant AcAlgE can play an important role in the degradation of acetylated alginate such as mucoidal acetylalginate in cystic fibrosis patient.

Enantioselective hydrolysis of racemic styrene oxide and its substituted derivatives using newly-isolated Sphingopyxis sp. exhibiting a novel epoxide hydrolase activity.

(S)-Styrene oxide, (S)-2-chlorostyrene oxide (CSO), (S)-3-CSO and (S)-4-CSO with 99.9 %ee were obtained with a yield of 20.6, 39.3, 28.7 and 26.8 % from 4 mM corresponding racemic substrates using 10 mg cells of a newly-isolated Sphingopyxis sp. at pH 8.0 and 25 °C in 1 ml 100 mM Tris/HCl buffer after 420, 100, 120 and 55 min, respectively. For racemic 2CSO, well-known for one of the racemates that is difficult to obtained in enantiomerically pure form, (S)-2-CSO with 99.9 %ee, 39.3 % yield (theoretical yield 50 %) and enantiomeric ratio of 42.1 was obtained. The newly-isolated strain can thus be used as whole-cell biocatalyst in the production of various (S)-CSO with a chlorine group at different positions.