Protein Electrophoretic Pattern of Two Chlorella Mutants with Low and High Content of Sulphur-Amino Acids

Two mutants of Chlorella vulgaris characterized by high (HS) or low (LS) content of sulphur-amino acids were compared to the wild strain with respect to their protein electrophoretic pattern. Three new bands, not present in the LS and wild strains, were found in the HS mutant. In algae grown in the presence of ³⁵S labelled sulphate two of these new bands showed a very high ³⁵S specific activity. Some electrophoretic bands common to LS, HS, and wild strain, were present in different proportions and showed different levels of specific radioactivity for each strain. Therefore, mutational events appear to have affected both the amino acid composition of single proteins and the relative amount of proteins with high and low sulphate content.     

Separation and simultaneous determination of nalidixic acid, hydroxynalidixic acid and carboxynalidixic acid in serum and urine by micellar electrokinetic capillary chromatography

Separation in capillary electrophoresis is governed by various factors, including buffer type, buffer concentration, pH, temperature, voltage and micelles. Through proper adjustment of these parameters, nalidixic acid and its two major metabolites, 7-hydroxynalidixic and 7-carboxynalidixic, could be separated by micellar electrokinetic capillary chromatography using an electrophoretic electrolyte consisting of 50 mM borate buffer (pH 9) containing 25 mM sodium dodecyl sulphate and 10% acetonitrile. A linear relationship between concentration and peak area for each compound was obtained in the concentration range 0.15–100 μg ml⁻¹, with a correlation coefficient greater than 0.999 and detection limits in the 0.2–0.7 ng ml⁻¹ range. Intra- and inter-day precision values of about 0.8–1.2% RSD (n=11) and 1.3–2.0% RSD (n=30), respectively, were obtained. The method has been applied to the analysis of nalidixic acid and its two major metabolites in serum and urine with limits of sensitivity lower than 0.8 ng ml⁻¹.     

Analyses of genetic variants of l-glycerol-3-phosphate dehydrogenase in Drosophila melanogaster by two-dimensional gel electrophoresis and immunoelectrophoresis

A protein spot corresponding to l-glycerol-3-phosphate dehydrogenase (α-GPDH, E.C. 1.1.1.8, NAD⁺ oxidoreductase) has been identified on a two-dimensional gel (isoelectric focusing-SDS gel) containing up to 150 stained protein spots from a crude Drosophila homogenate. Preliminary identification of the α-GPDH spot was made by including a suitable amount of purified Drosophila α-GPDH in crude fly homogenates prior to electrophoresis and observing an intensity enhancement of the corresponding protein spot on the gels. When three purified electrophoretic variants (slow, fast, and ultrafast) were mixed and analyzed by two-dimensional gel electrophoresis, horizontal displacements of the three protein spots were observed. Immunoprecipitation of the enzyme prior to electrophoresis and gene mapping further confirmed the identity of the α-GPDH protein spot. The α-GPDH spot can also be detected by autoradiography of a two-dimensional gel from a single fly extract, where it has been estimated to constitute 0.5–1% of the total soluble protein. Mutants which express no apparent α-GPDH activity were analyzed by two-dimensional gels and immunoelectrophoresis in an attempt to identify and characterize the inactive proteins. It is suggested that these techniques provide a powerful tool for the analysis of CRM⁺-null activity mutants of a specific gene-enzyme system.     

Relaxed mutants ofSerratia marcescens SM-6. Biochemical traits and relevance of therel + allele for the formation of exoenzymes

Serratia marcescens SM-6 when starved for a required amino acid stops synthesizing protein and RNA and accumulates two nucleotides which cochromatograph with ppGpp and pppGpp. These features are characteristic of bacterial strains with stringent RNA control (rel ⁺). Two independent mutants were isolated which resemble relaxed (relA) mutants ofEscherichia coli; they continue to synthesize RNA and accumulate neither ppGpp nor pppGpp when deprived of the required amino acid. The extracellular enzyme activities (nuclease, protease, lipase) of the relaxed mutants are about the same as those of the parental stringent strain when studied under standard growth conditions. Exoenzyme-deficient (nuc; prt) and exoenzyme-hyperproducing (nuc ^su) mutants were isolated from both stringent and relaxed strains ofS. marcences SM-6 and no change of the cellular ability to form ppGpp and pppGpp could be observed. From these results it appears that the formation of exoenzymes ofS. marcescens SM-6 is independent of stringent/relaxed RNA control.Abbreviations cpd cyclic nucleotide phosphodiesterase deficient - nuc nuclease deficient - nuc ^su nuclease hyperproducing - prt protease deficient - rel relaxed control - spo ppGpp deficient (spot less) - ppGpp guanosine tetraphosphate - pppGpp guanosine pentaphosphate - TCA trichloroacetic acid - OD optical density - EU enzyme units     

Characterization of recombinant endo-1,4-β-xylanase of Bacillus halodurans C-125 and rational identification of hot spot amino acid residues responsible for enhancing thermostability by an in-silico approach

Increased demand of enzymes for industrial use has led the scientists towards protein engineering techniques. In different protein engineering strategies, rational approach has emerged as the most efficient method utilizing bioinformatics tools to produce enzymes with desired reaction kinetics; physiochemical (temperature, pH, half life, etc) and biological (selectivity, specificity, etc.) characteristics. Xylanase is one of the widely used enzymes in paper and food industry to degrade xylan component present in plant pulp. In this study endo 1,4-β-xylanase (Xyl-11A) from Bacillus halodurans C-125 was cloned in pET-22b (+) vector and expressed in Escherichia coli BL21 (DE3) expression strain. The enzyme had Michaelis constant K_m of 1.32 mg ml^?1 birchwoodxylan (soluble form) and maximum reaction velocity (V_max) 73.53 mmol min^?1 mg^?1 with an optimum temperature of 75 °C and pH 9.0. The thermostability analysis showed that enzyme retained more than 80% of its residual activity when incubated at 75 °C for 2 h. In addition, to increase Xyl-11A thermostability, an in-silico analysis was performedto identify the hot spot amino acid residues. Consensus-based amino acid substitution was applied to evaluate multiple sequence alignment of homologs and identified 20 amino acids positions by following Jensen-Shnnon Divergence method. 3D models of 20 selected mutants were analyzed for conformational transition in protein structures by using NMSim server. Two selected mutants T6K and I17M of Xyl-11A retained 40, 60% residual activity respectively, at 85 °C for 120 min as compared to wild type enzyme which retained 37% initial activity under same conditions, confirming the enhanced thermostability of mutants. The present study showed a good approach for the identification of promising amino acid residues responsible for enhancing the thermostability of enzymes of industrial importance.

     

Comparison of purified acid phosphatase allozymes inDrosophila virilis: Differences in carbohydrate content and composition of the allozymes

Three acid phosphatase (EC 3.1.3.2) allozymes (ACPH¹, ACPH², and ACPH⁴) ofDrosophila virilis show different activities as measured by electrophoretic techniques. Recently, it was suggested that these differences are attributable to the variable ability of the allozymes to be incorporated into lysosomes (Narise, S.,Genet. Res. Cambr., 45:143, 1985). Immunoelectrophoresis demonstrated that the activity differences between these electrophoretic variants coincided with differences in the amount of the enzyme protein in soluble fractions but not in whole cell-free extracts. These results support the idea that acid phosphatase allozymes inD. virilis are cell-localization variants. We examined the problem by structural analysis of both the protein and the carbohydrate moieties of these allozyme glycoproteins, since lysosomal enzymes are known to become localized in lysosomes through their carbohydrate moieties. The three ACPH allozymes were purified to homogeneity from their respective homozygotes and compared with respect to amino acid composition and carbohydrate content and composition. Amino acid compositions were similar, while content and compositions of neutral sugars were significantly different. The neutral sugar content of ACPH¹ was 9.2%; that of ACPH², 21.0%; and that of ACPH⁴, 7.3%. A trace of hexosamines, but noN-acetylneuraminic acid, was found in the ACPH allozymes. Isoelectric points varied corresponding to their electrophoretic mobilities, which were not changed by treatment with alkaline phosphatase and neuraminidase.     

Characterisation of new symbiotic Medicago truncatula (Gaertn.) mutants, and phenotypic or genotypic complementary information on previously described mutants

From a pool of Medicago truncatula mutants—obtained by gamma-irradiation or ethyl methanesulfonate mutagenesis—impaired in symbiosis with the N-fixing bacterium Sinorhizobium meliloti, new mutants are described and genetically analysed, and for already reported mutants, complementary data are given on their phenotypic and genetic analysis. Phenotypic data relate to nodulation and mycorrhizal phenotypes. Among the five new mutants, three were classified as [Nod⁺ Fix^– Myc⁺] and the mutations were ascribed to two loci, Mtsym20 (TRV43, TRV54) and Mtsym21 (TRV49). For the two other new mutants, one was classified as [Nod^–/+ Myc⁺] with a mutation ascribed to gene Mtsym15 (TRV48), and the other as [Nod^– Myc^-/+] with a mutation ascribed to gene Mtsym16 (TRV58). Genetic analysis of three previously described mutants has shown that [Nod^–/+ Myc⁺] TR74 mutant can be ascribed to gene Mtsym14, and that [Nod^–/+ Myc^–/+] TR89 and TRV9 mutants are ascribed to gene Mtsym2 (dmi2). Using a detailed analysis of mycorrhizal phenotype, we have observed a delayed typical arbuscular mycorrhizal formation on some mutants that present thick lens-shaped appressoria. This phenotype was called [Myc^–/+] and mutants TR25, TR26, TR89, TRV9, P1 and Y6 were reclassified as [Myc^–/+]. Mutant P1 was reclassified as [Nod^–/+] because of a late nodulation observed on roots of this mutant.     

Genetic analysis of two weak dormancy mutants derived from strong seed dormancy wild type rice N22 (Oryza sativa)

Two weak dormancy mutants, designated Q4359 and Q4646, were obtained from the rice cultivar N22 after treatment with 400 Gy ⁶⁰Co gamma‐radiation. Compared to the N22 cultivar, the dormancy of the mutant seeds was more readily broken when exposed to a period of room temperature storage. The mutants also showed a reduced level of sensitivity to abscisic acid compared to the N22 cultivar, although Q4359 was more insensitive than Q4646. A genetic analysis indicated that in both mutants, the reduced dormancy trait was caused by a single recessive allele of a nuclear gene, but that the mutated locus was different in each case. The results of quantitative trait locus (QTL) mapping, based on the F₂ population from Q4359 x Nanjing35, suggested that Q4359 lacks the QTL qSdn‐1 and carries a novel allele at QTL qSdn‐9, while a similar analysis of the Q4646 x Nanjing35 F₂ population suggested that Q4646 lacks QTL qSdn‐5, both qSdn‐1 and qSdn‐5 are major effect seed dormancy QTL in N22. Therefore, these two mutants were helpful to understand the mechanism of seed dormancy in N22.     

Semi-rational site-directed mutagenesis of phyI1s from <Emphasis Type="Italic">Aspergillus niger</Emphasis> 113 at two residue to improve its phytase activity

Through alignment of amino acid sequences among different phytases, we found that the amino acid at residues 53 and 91 vary broadly. To prove that the amino acid at residues 53 and 91 were related to phytase specific activity, two single mutant phyI1s Q53R and K91D were obtained by site-directed mutagenesis strategy. None of the single amino acid residues in the two mutants was in a position reported to be important for catalysis or substrate binding. Kinetic analysis of the phytase activity of the two mutants (Q53R and K91D) indicated that the mutants were attributed to 2.2- and 1.5-fold increased specific activity, and a 1.47- and 1.16-fold increased affinity for sodium phytate. In addition, the overall catalytic efficiency (k _cat/K _m) of the two mutants was improved 4.08- and 2.84-fold compared to that of the wild type. Such mutants will be instrumental for the structure–function study of the enzyme and for industrial application.     

Genetics and biochemistry of the peptidoglycan-associated proteins b and c of Escherichia coli K12.

Summary To collect information on synthesis and regulation of the peptidoglycan-associated pore-forming outer membrane proteins b and c, mutants resistant to phages Mel and TuIa were analyzed. Genetic analysis showed three linkage groups, corresponding with the genes tolF (phenotype b^-c⁺), meo A (phenotype b⁺c^-) and ompB (phenotypes b^-c^-, b^- c⁺, b⁺⁺ c^- and b⁺⁺ c^±). It has recently been described that also a b⁺ c^- phenotype can occur in the latter linkage group [Chai, T., Foulds, J., J. Bacteriol. 130, 781–786 (1977)]. Among ompB (b^- c⁺)/meoA (b⁺ c^-) double mutants strains were found with the b⁺ c^- phenotype, showing that ompB is not the structural gene for protein b. Studies on purified proteins b and c showed profound differences between the two proteins with respect to the electrophoretic mobility of fragments obtained by treatment with cyanogen bromide, trypsin and chymotrypsin. The amino acid in position three of the amino-termini of proteins b and c, isolated from isogenic strains, were identified as isoleucine and valine respectively. Both the genetic and biochemical results are consistent with a model recently published [Ichihara, S., Mizushima, S., J. Biochem. (Japan) 83, 1095–1100 (1978)] which predicts that tolF and meoA are the structural genes for the proteins b and c respectively and that ompB is a regulatory gene whose product regulates the levels of both proteins.     

Discovery of a genetic polymorphism of human plasma protein C inhibitor (PCI): genetic survey utilizing isoelectric focusing followed by immunoblotting,immunological and biochemical characterization

Summary The objectives of this study were to determine the genetic basis of the electrophoretic differences of human plasma protein C inhibitors (PCI) from 977 individuals. Three discrete antibodies were produced against the PCI purified from human plasma and peptides that corresponded to the N-terminal 15 amino acid residues and the C-terminal 15 residues of human PCI, the chemical structures of which were determined by cDNA sequence analysis. The combined techniques of polyacrylamide gel isoelectric focusing and immunoblotting with these three different antibodies resolved the plasma PCI into several isoprotein bands, with a pH range of 6–7. These PCI isoproteins, however, were not stained by anti-human kallikrein, anti-human protein C or anti-human urokinase antibodies. Therefore, each of the PCI bands, which were detected by immunoblotting with the anti-PCI antibody and the two different anti-peptide antibodies, were derived from free PCI, and not an inactive PCI species. Two common phenotypes, designated PCI 1 and 1–2, were recognized, and family studies showed that they represented homozygosity or heterozygosity for two autosomal codominant alleles, PCI ^*1 and PCI ^*2. A population study of plasma samples collected from 977 Japanese individuals indicated that the frequencies of the PCI ^*1 and PCI ^*2 alleles were 0.988 and 0.012, respectively.     

低H~+_-ATPase活性植物乳杆菌突变菌筛选及基因表达的相对定量分析

【目的】筛选H~+_-ATPase活性降低的植物乳杆菌突变菌,比较其与亲本菌基因表达水平的差异,进一步探索H~+_-ATPase的调控机制。【方法】利用硫酸新霉素诱变、筛选突变菌,并对亲本菌(ZUST)和突变菌(ZUST-1、ZUST-2)进行生长、产酸能力及H~+_-ATPase活性的测定。分别提取亲本菌和突变菌的基因组DNA,扩增H~+_-ATPase全部编码基因并测序。通过荧光定量PCR对H~+_-ATPase全部编码基因进行相对定量分析。【结果】突变菌的生长和产酸能力均低于亲本菌,突变菌ZUST-1和ZUST-2的H~+_-ATPase活性比亲本菌分别降低了10.1%和28.8%。突变菌ZUST-1和ZUST-2的atp A基因均有22个位点发生突变,而ZUST-2的atp C基因有6个位点发生突变。突变菌ZUST-1和ZUST-2的atp A在对数期基因表达水平分别比亲本菌ZUST下调了41.1%和35.7%,在稳定期分别下调了43.6%和14.2%;ZUST-1的atp C基因在对数期的表达水平比ZUST略高,在稳定期比ZUST上调了30%,而ZUST-2的atp C基因未表达。【结论】突变菌H~+_-ATPase活性减弱会导致其全部编码基因在稳定期表达水平上调(除ZUST-2的atp C不表达外),而且atp A和atp C基因突变导致的基因表达水平的差异是影响H~+_-ATPase活性的主要因素,此研究结果为进一步研究植物乳杆菌中H~+_-ATPase的调控机制奠定了基础。     

Electrophoretically detectable mutations induced in CHO cells by varying doses of ultraviolet radiation

Chinese hamster ovary (CHO) cells were subjected to ultraviolet radiation (UV) at doses resulting in 100% (no irradiation), 50–30%, 20–10% and ≈1% survival. 2 divisions after UV exposure surviving cells were cloned and clones expanded for electrophoretic analysis of the products of ≈40 enzyme loci. 4 different classes of variants (electrophoretic shifts, nulls, enzyme re-expression and enzyme modification) were detected in 29 of 1329 clones analyzed and proven mutants by subclone analysis. The frequency of mutants in the irradiated groups (28/38391 loci screened or 7.3 × 10^?4) was significantly higher than controls. The frequency of shift mutants at 10–20% survival was higher than shifts at 30–50% survival and was significantly higher than shifts at ≈1% survival. The frequency of nulls increased with dose. 12 of the 28 mutants obtained in the irradiated groups were at only 3 of the mean 41 loci screened/clone. The results indicated that shift mutants could be detected more efficiently than nulls at lower dose and that loci varied widely with respect to their susceptibility to UV mutagenesis. Multiple null mutants at 2 loci, isocitrate dehydrogenase 2 and hexokinase 2, indicated they may be hemizygous in CHO cells.     

Analysis of Tomato spotted wilt virus NSs protein indicates the importance of the N‐terminal domain for avirulence and RNA silencing suppression

Recently, Tomato spotted wilt virus (TSWV) nonstructural protein NSs has been identified unambiguously as an avirulence (Avr) determinant for Tomato spotted wilt (Tsw)‐based resistance. The observation that NSs from two natural resistance‐breaking isolates had lost RNA silencing suppressor (RSS) activity and Avr suggested a link between the two functions. To test this, a large set of NSs mutants was generated by alanine substitutions in NSs from resistance‐inducing wild‐type strains (NSs^RI), amino acid reversions in NSs from resistance‐breaking strains (NSs^RB), domain deletions and swapping. Testing these mutants for their ability to suppress green fluorescent protein (GFP) silencing and to trigger a Tsw‐mediated hypersensitive response (HR) revealed that the two functions can be separated. Changes in the N‐terminal domain were found to be detrimental for both activities and indicated the importance of this domain, additionally supported by domain swapping between NSs^RI and NSs^RB. Swapping domains between the closely related Tospovirus Groundnut ringspot virus (GRSV) NSs and TSWV NSs^RI showed that Avr functionality could not simply be transferred between species. Although deletion of the C‐terminal domain rendered NSs completely dysfunctional, only a few single‐amino‐acid mutations in the C‐terminus affected both functions. Mutation of a GW/WG motif (position 17/18) rendered NSs completely dysfunctional for RSS and Avr activity, and indicated a putative interaction between NSs and Argonaute 1 (AGO1), and its importance in TSWV virulence and viral counter defence against RNA interference.     

Rapid selection of mutants of <Emphasis Type="Italic">Chlamydomonas reinhardtii</Emphasis> for carbohydrate and fatty acid metabolism by fourier transform infrared spectroscopy and gas chromatography combined with multivariate analysis

Transgene-tagged mutants of Chlamydomonas reinhardtii were generated by random insertional mutagenesis for screening of mutants of carbohydrate and fatty acid metabolism. Approximately 2,500 insertion mutants tagged with the aph7″ gene were produced from one mutagenesis in three weeks. To establish a rapid screening system for numerous insertional lines, whole cell extracts of 100 insertional lines were subjected to Fourier transform infrared spectroscopy (FT-IR) and gas chromatography (GC) analysis combined with multivariate analysis. Mutant lines 28, 67, and 90 showed dramatic differences in the carbohydrate (1,000∼1,200 cm⁻¹) and amide (1,500∼1,700 cm⁻¹) regions of the FT-IR spectrum compared to wild type strain CC-124. Separate GC analysis also showed that 16:0 iso, palmitic acid (16:0), and oleic acid (18:1) were the major fatty acids in the wild type strain. In mutant 80, the relative content ratio of 16:0 iso in total fatty acids was significantly lower than in wild type, whereas the ratios of palmitic acid and oleic acid to 16:0 iso were higher. In mutant 95, the ratio of 16:0 iso to total fatty acids was increased, whereas ratios of palmitic acid and oleic acid to 16:0 iso were decreased. In particular, mutant 57 showed remarkably different fatty acid patterns with novel peaks of long-chain fatty acids having more than 20 carbon atoms. The results of this study show that FT-IR and GC combined with multivariate analysis enable rapid selection of mutants of carbohydrate and fatty acid metabolism in C. reinhardtii.     

Isolation and characterization of further cis- and trans-acting regulatory elements involved in the synthesis of glucose-repressible alcohol dehydrogenase (ADHII) in Saccharomyces cerevisiae.

Summary Starting with yeast cells lacking the constitutive alcohol dehydrogenase activity (ADHI), mutants with partially glucose-insensitive formation of ADHII were isolated. Genetic analysis showed that four mutants (designated ADR3 ^c) were linked to the ADHII-structural gene, ADR2, and were cis-dominant. On derepression, two of them produced elevated ADHII-levels, indicating a promotor function of the altered controlling site. The other ADR3 ^c-mutant alleles affected the ADHII-subunit association in diploids carrying two electrophoretically distinct alleles of the structural gene ADR2. Twelve semidominant constitutive mutants could be attributed to gene ADR1 (ADR1 ^c-alleles) previously identified by recessive mutants with blocked derepression. This suggested a positive regulatory role of the ADR1 gene product on the expression of the ADHII-structural gene. A pleiotropic mutation ccr1 (Ciriacy, 1977) was epistatic over glucose-resistant ADHII-formation caused by ADR1 ^c-alleles. From this it was concluded that CCR1 specifies for a product co-activating the structural gene or modifying the ADR1-gene product. A further regulatory element (gene designation ADR4) not linked to the structural gene could be identified upon isolation of recessive constitutive mutants adr4 from a ccr1 ADR1 ^c-double mutant.     

Mutations in two amino acids in phyI1s from <Emphasis Type="Italic">Aspergillus niger</Emphasis> 113 improve its phytase activity

We applied in vitro mutagenesis and colony screening, using the wild type phyI1s gene from Aspergillus niger 113 as the template, and obtained two mutant phyI1s (gene products) after one round of screening. The two mutants had mutations at two nucleic acid sites, resulting in changes in two amino acids: K41E, E121F. None of the amino acid substitutions in the two mutants was in a position reported to be important for catalysis or substrate binding. Kinetic analysis of the phytase activity of the two mutants indicated that the substitutions gave rise to 2.5- and 3.1-fold increased specific activity, and a 1.78- and 3.24-fold reduced affinity for sodium phytate. In addition, the overall catalytic efficiency (k _cat/K _m) of the two mutants was changed by 0.52-fold and 0.68-fold compared to that of the wild type. Such mutants will be instrumental for the structure–function study of the enzyme and for industrial application.     

Undermodification in the first position of the anticodon of supG-tRNA reduces translational efficiency

Summary Two mutants of Escherichia coli, trmC1 and trmC2, which are both defective in the synthesis of 5-methylaminomethyl-2-thiouridine (mnm⁵s²U) were utilized to study the function of this complex modified nucleoside. Transfer RNAs specific for glutamine, glutamic acid and lysine as well as a specific ochre suppressor derived from lysine tRNA (tRNA _UAA ^lys encoded by the supG allele), contain this modified nucleoside at position 34 (the wobble position). It was found that two different undermodified derivatives of mnm⁵s²U were present in the two trmC mutants, which suggests that the two mutations affect two different enzymatic activities. Using the lacI-Z fusion system (Miller and Albertini 1983), we found that the efficiency of supG-mediated suppression was reduced to 30%–90% of the wild-type value in the trmC mutants. The modificationdeficient supG-tRNA in the mutants showed a higher sensitivity to codon context than the normal tRNA _UAA ^lys .     

Fermentation of 1,3-propanediol by a lactate deficient mutant of Klebsiella oxytoca under microaerobic conditions

Klebsiella oxytoca M5al is an excellent 1,3-propanediol (1,3-PD) producer, but too much lactic acid yielded greatly lessened the fermentation efficiency for 1,3-PD. To counteract the disadvantage, four lactate deficient mutants were obtained by knocking out the ldhA gene of lactate dehydrogenase (LDH) of K. oxytoca M5al. The LDH activities of the four mutants were from 3.85 to 6.92% of the parental strain. The fed-batch fermentation of 1,3-PD by mutant LDH3, whose LDH activity is the lowest, was studied. The results showed that higher 1,3-PD concentration, productivity, and molar conversion rate from glycerol to 1,3-PD can be gained than those of the wild type strain and no lactic acid is produced under both anaerobic and microaerobic conditions. Sucrose fed during the fermentation increased the conversion and sucrose added at the beginning increased the productivity. In fed-batch fermentation with sucrose as cosubstrate under microaerobic conditions, the 1,3-PD concentration, conversion, and productivity were improved significantly to 83.56 g l⁻¹, 0.62 mol mol⁻¹, and 1.61 g l⁻¹ h⁻¹, respectively. Furthermore, 60.11 g l⁻¹ 2,3-butanediol was also formed as major byproduct in the broth.