Kegg Pathway: Butanoate metabolism

Curr Med Chem. 2009; 16(28): 3686-700
Quash G, Fournet G

Methionine, in addition to its role in protein synthesis, participates in 3 important cellular functions: as AdoMet in transmethylation; as decarboxylated-AdoMet in aminopropylation; as homocysteine its demethylated form, in trans-sulphuration. Here we provide evidence from the literature and from our own work for a fourth role for its oxoacid: 4-methylthio-2-oxo-Butanoate (MTOB) in apoptosis [28,29]. MTOB enters 2 pathways: (a) transamination by glutamine-transaminase K to methionine[13,14].(b)oxidative decarboxylation by the mitochondrial Branched-Chain-Oxo-Acid-Dehydrogenase-Complex to methional and finally to methylthiopropanoyl CoA (MTPCoA) [26,27]. Some of the methional formed after MTOB decarboxylation leaks into the cytoplasm as free methional [29]. Exogenous methional induces apoptosis in normal and cancer cells in culture [28, 29] but not in those overexpressing the antiapoptotic gene bcl2 [30]. In physiologically-induced apoptosis e.g; trophic factor (IL3) withdrawal, methional leakage is decreased [29] suggesting that MTPCoA is also involved in apoptosis. Both methional and MTPCoA give rise to metabolites that may act as cross-linking agents. In the case of methional, the CH3-S moiety is lost and malondialdehyde (MDA) is formed when methional is subjected to ( )OH attack [29]. MDA generated in situ from 1,3-propanediol, induces DNA-protein cross-linking [41].With regard to MTPCoA, it is metabolized to malonic semialdehyde CoA (MASACoA) with loss of the CH3-S moiety [48,49]. The capacity of MASACoA to form cross-links has not yet been established experimentally, but it could be a substrate for one of the histone acyl transferases [50, 51] and so form amides via the CoA at one end and imines by its CHO group at the other, with amino groups on proteins. Chromatin cross-linking/condensation is one of the hall-marks of apoptosis [40]. Methional, MDA and other apoptogenic aldehydes like 4-hydroxy-2-nonenal are oxidized by ALDHs to non-apoptogenic carboxylic acids [29,44, 45,68] but retain their apoptotic activity when the ALDHs are inhibited [98,110]. MASACoA would also lose its cross-linking capacity if its CoA moiety were putatively hydrolysed by ALDHs and/or acylCoA thioesterases [56,58,88,89]. ALDH inhibitors that control cellular MDA and possibly MASACoA homeostasis are cited as examples of targeted therapeutic approaches in chemoresistant cancers [62,84,97,98,110].

J Proteome Res. 2009 Apr; 8(4): 1623-30
Bao Y, Zhao T, Wang X, Qiu Y, Su M, Jia W, Jia W

The pathological development and the drug intervention of type 2 diabetes mellitus (T2DM) involve altered expression of downstream low molecular weight metabolites including lipids and amino acids, and carbohydrates such as glucose. Currently, a small number of markers used for clinical assessment of T2DM treatment may be insufficient to reflect global variations in pathophysiology. In this study, a metabonomic study was performed to determine metabolic variations associated with T2DM and the drug treatments on 74 patients who were newly diagnosed with T2DM and received a 48 week treatment of a single drug, repaglinide, metformin or rosiglitazone. Fasting overnight and 2 h postprandial blood serum of patients were collected at 24 and 48 weeks to monitor the biochemical indices (FPG, 2hPG, HbA1c, etc.). Gas chromatography/mass spectrometer coupled with multivariate statistical analysis was used to identify the alteration of global serum metabolites associated with T2DM as compared to healthy controls and responses to drug treatment. Significantly altered serum metabolites in diabetic subjects include increased valine, maltose, glutamate, urate, Butanoate and long-chain fatty acid (C16:0, C18:1, C18:0, octadecanoate and arachidonate), and decreased glucuronolactone, lysine and lactate. All of the three treatments were able to down-regulate the high level of glutamate to a lower level in serum of T2DM patients, but rosiglitazone treatment was able to reverse more abnormal levels of metabolites, such as valine, lysine, glucuronolactone, C16:0, C18:1, urate, and octadecanoate, suggesting that it is more efficient to alter the metabolism of T2DM patients than the other two drugs.

Arch Virol. 2009; 154(9): 1539-44
Colombo L, Piovesan P, Ghirardi O, Salmona M, Forloni G

On the basis of the structural homologies between ST1859 (1[(2-hydroxy-1-naphtyl)methyl]-2-naphthol) and the anti-prion agents and its anti-amyloidogenic activity, we tested whether this molecule altered the biochemical properties of aggregates formed in vitro by synthetic prion peptides and affected prion infectivity in experimental scrapie. Co-incubation of ST1859 with the peptides PrP 106-126 and PrP 82-146 reduced their fibrillogenic capacity and their resistance to digestion with protease K. Hamsters inoculated with the ST1859-treated homogenate showed a significant delay in the onset of clinical signs of disease and longer survival. Survival was also significantly longer in infected hamsters treated peripherally with ST1859 for the whole post-inoculation period until the onset of clinical symptoms. Similar results were found with the analogue ST1745. Our data indicate that ST1859 reduces prion infectivity and can exert a therapeutic effect in experimental scrapie.

Arch Latinoam Nutr. 2009 Mar; 59(1): 88-94
Acosta O, Pérez AM, Vaillant F

Naranjilla (Solanum quitoense Lam.) is a native fruit of the Andes, cultivated and consumed mainly in Ecuador, Colombia, and Central America. Because of its pleasant aroma and attractive color, it has high potential as an ingredient of products such as juices, nectars, and jams. The main characteristics of mature naranjilla fruits cultivated in Costa Rica were assessed, including sugar content, total titratable acidity, total soluble solids, oxygen radical absorbance capacity (H-ORAC), and total polyphenolic and ascorbic acid content. Carotenoid and volatile compound identification was also done. The samples showed sucrose, glucose, and fructose content of 1.6 +/- 0.3, 0.68 +/- 0.05, and 0.7 +/- 0.1 g/100 g, respectively. Total titratable acidity was 2.63 +/- 0.07 g citric acid equivalent / 100 g and total soluble solids amounted to 9.1 +/- 0.5 degrees Brix. H-ORAC value was 17 +/- 1 micromol Trolox equivalent /g, total polyphenolic content was 48 +/- 3 mg gallic acid equivalent /100 g and ascorbic acid content was 12.5 +/- 0.0 mg/100 g. Carotenoid content of the whole fruit and pulp was 33.3 +/- 0.6 and 7.2 +/- 0.3 microg/g, respectively. The predominant carotenoid among the compounds identified in the whole fruit was beta-carotene. Ten volatile compounds were identified in naranjillapulp, the predominant being methyl Butanoate. The chemical composition of naranjilla cultivated in Costa Rica does not seem to differ from that previously reported in studies at different locations.

Drug Metab Lett. 2008 Dec; 2(4): 275-9
Morris AP, Brain KR, Heard CM

In probing enhancement of the transdermal delivery of the anti-psychotic drug haloperidol, five prodrugs (ethanoate, propanoate, Butanoate, octanoate and decanoate) were synthesised and their relative rates of hydrolysis determined in the presence of porcine liver esterase (PLE), a model for cutaneous esterases. (1)H NMR, MS and elemental analysis confirmed the successful synthesis of each prodrug in high purity, and each was found to hydrolyse in the presence of PLE with the hydrolytic rate reaching a maximum with haloperidol octanoate (C8) at 2.31 +/- 0.06 nmol ml(-1) h(-1) (p < 0.001).

J Agric Food Chem. 2009 Apr 8; 57(7): 2875-81
Zhang B, Yin XR, Li X, Yang SL, Ferguson IB, Chen KS

During postharvest ripening of kiwifruit [ Actinidia deliciosa (A. Chev.) C.F. Liang et A.R. Ferguson var. deliciosa cv. Bruno] at 20 degrees C, six lipoxygenase (LOX) genes exhibited different expression patterns. AdLox1 and AdLox5 were up-regulated during ripening, and transcript accumulation was delayed by 1-methylcyclopropene (1-MCP), whereas AdLox2, AdLox3, AdLox4, and AdLox6 were down-regulated with ripening. Levels of two volatiles arising from the LOX pathway, that is, n-hexanal and (E)-2-hexenal, were highest after harvest and declined during ripening at 20 degrees C, whereas the characteristic kiwifruit esters ethyl and methyl Butanoate levels increased late in the ripening process. Individual fatty acid concentrations underwent little change during ripening, with linoleic (LA) and linolenic (LeA) acids constituting about 40% of the total. Application of LA and LeA to kiwifruit flesh disks promoted LOX activity and n-hexanal and (E)-2-hexenal generation, whereas inhibitors of LOX, n-propyl gallate (n-PG) and nordihydroguariaretic acid (NDGA), caused a parallel reduction in enzyme activity and in the production of C6 aldehydes. The six LOX genes showed different sensitivities to the LOX substrates and inhibitors. The ethylene up-regulated genes AdLox1 and AdLox5 were induced by LA and LeA and inhibited by n-PG and NDGA. Of the LOX genes that were down-regulated by ethylene, only AdLox4 and AdLox6 were stimulated in response to the substrates and retarded by the inhibitors. The possible roles of the six LOX genes in kiwifruit ripening and aroma development are discussed.

J Agric Food Chem. 2009 Feb 11; 57(3): 974-80
Almenar E, Hernández-Muñoz P, Gavara R

The effect of chitosan coating on the evolution of several volatile compounds relevant to the strawberry ( Fragaria x ananassa cv. Camarosa) aroma profile has been investigated. Strawberries dipped in chitosan acetate solution at 1 or 1.5% (w/w) and uncoated controls were stored at 10 degrees C for 1 week. Significant differences in aroma profile between coated and uncoated samples were observed. Most importantly, the buildup of the off-flavors acetaldehyde and ethanol was largely delayed in coated berries. With regard to the effect of chitosan on ester evolution, the levels of ethyl Butanoate and ethyl hexanoate, important contributors to strawberry aroma related to fruity and sweet notes, were found to be enhanced in coated fruit. Acetate esters also increased during storage but more markedly in uncoated strawberries. These results show the potential of chitosan coatings in maintaining strawberry flavor during storage, something difficult to achieve with current conservation technologies. Moreover, differences in results for different coating concentrations are reported.

J Proteome Res. 2009 Jan 22;
Bao Y, Zhao T, Wang X, Qiu Y, Su M, Jia W, Jia W

The pathological development and the drug intervention of type 2 diabetes mellitus (T2DM) involve altered expression of downstream low molecular weight metabolites including lipids and amino acids, and carbohydrates such as glucose. Currently, a small number of markers used for clinical assessment of T2DM treatment may be insufficient to reflect global variations in pathophysiology. In this study, a metabonomic study was performed to determine metabolic variations associated with T2DM and the drug treatments on 74 patients who were newly diagnosed with T2DM and received a 48 week treatment of a single drug, repaglinide, metformin or rosiglitazone. Fasting overnight and 2 h postprandial blood serum of patients were collected at 24 and 48 weeks to monitor the biochemical indices (FPG, 2hPG, HbA(1c), etc.). Gas chromatography/mass spectrometer coupled with multivariate statistical analysis was used to identify the alteration of global serum metabolites associated with T2DM as compared to healthy controls and responses to drug treatment. Significantly altered serum metabolites in diabetic subjects include increased valine, maltose, glutamate, urate, Butanoate and long-chain fatty acid (C16:0, C18:1, C18:0, octadecanoate and arachidonate), and decreased glucuronolactone, lysine and lactate. All of the three treatments were able to down-regulate the high level of glutamate to a lower level in serum of T2DM patients, but rosiglitazone treatment was able to reverse more abnormal levels of metabolites, such as valine, lysine, glucuronolactone, C16:0, C18:1, urate, and octadecanoate, suggesting that it is more efficient to alter the metabolism of T2DM patients than the other two drugs.

Riv Biol. 2008 Jan-Apr; 101(1): 67-80
Ding D, Ding Y, Cai Y, Chen S, Xu W

The objective of this article is to obtain a more detailed insight into poly-beta-hydroxybutyrate (PHB) metabolism through network-based metabolic pathway analysis. We employ extreme pathways to perform this study, because calculating and interpreting extreme pathways is a promising way for pathway analysis and metabolic engineering. After giving an in silico model of Butanoate metabolism of Bacillus thuringiensis 97-27 (btk), extreme pathways were calculated and classified. Furthermore, the type I and II extreme pathways were further classified and analyzed in detail based on their structure and functional capabilities. Besides "historical" biochemical pathways, the results also suggest that there are some novel pathways.

J Chem Ecol. 2008 Jun; 34(6): 742-7
Tschuch G, Lindemann P, Moritz G

Adults and larvae of the thrips Callococcithrips fuscipennis (Moulton) (Thysanoptera: Tubulifera: Phlaeothripidae) live in the sticky wax masses of adult females of the felt scale insect Callococcus acaciae (Maskell) (Sternorrhyncha: Coccoidea: Eriococcidae). The scale is sessile and feeds on Kunzea shrubs (Myrtales: Myrtaceae). If stressed, the thrips produce droplets of secretions. The mixture contains pentadecane, tridecane, two monoterpenoids, hexadecyl Butanoate, and smaller amounts of 15 other esters of long-chain unbranched alcohols identified as acetates, Butanoates, hexanoates, and octanoates. The monoterpenoids are dolichodial, an iridoid, and an unknown substance with a mass spectrum very similar to that of anisomorphal and peruphasmal, diastereomers of dolichodial, but with a different retention time. Iridoids, Butanoates, hexanoates, and octanoates have not been previously identified in Thysanoptera.

J Org Chem. 2008 May 16; 73(10): 3875-84
Taniguchi T, Fukuba TA, Nakatsuka S, Hayase S, Kawatsura M, Uno H, Itoh T

Candida antarctica lipase B (CAL-B) is one the most frequently used enzymes in organic synthesis for the preparation of optically active alcohols. However, it has not been used for the optical resolution of (+/-)-2,2'-binaphthol. We established an efficient linker-oriented design of 2,2'-binaphthol derivatives that is appropriate for optical resolution using CAL-B-catalyzed hydrolysis reaction. Methyl 4-(1-(6-bromo-2-methoxymethoxynaphthalen-1-yl)-6-bromonaphthalen-2-yloxy)Butanoate was hydrolyzed by CAL-B to afford a corresponding acid with excellent enantioselectivity ( E > 200). Two types of optically active binaphthol derivatives, 1-(2-hydroxy-6-(naphthalen-1-yl)naphthalen-1-yl)-6-(naphthalen-1-yl)naphthalen-2-ol and 6-butyl-1-(6-butyl-2-hydroxynaphthalen-1-yl)naphthalen-2-ol, were prepared by this chemo-enzymatic reaction protocol and were used as chiral templates for symmetric reactions.

Mol Microbiol. 2008 Apr; 68(1): 152-72
Zuther K, Mayser P, Hettwer U, Wu W, Spiteller P, Kindler BL, Karlovsky P, Basse CW, Schirawski J

Tryptophan is a precursor for many biologically active secondary metabolites. We have investigated the origin of indole pigments first described in the pityriasis versicolor-associated fungus Malassezia furfur. Some of the identified indole pigments have properties potentially explaining characteristics of the disease. As M. furfur is not amenable to genetic manipulation, we used Ustilago maydis to investigate the pathway leading to pigment production from tryptophan. We show by high-performance liquid chromatography, mass spectrometry and nuclear magnetic resonance analysis that the compounds produced by U. maydis include those putatively involved in the etiology of pityriasis versicolor. Using a reverse genetics approach, we demonstrate that the tryptophan aminotransferase Tam1 catalyses pigment biosynthesis by conversion of tryptophan into indolepyruvate. A forward genetics approach led to the identification of mutants incapable of producing the pigments. These mutants were affected in the sir1 gene, presumably encoding a sulphite reductase. In vitro experiments with purified Tam1 showed that 2-oxo 4-methylthio Butanoate serves as a substrate linking tryptophan deamination to sulphur metabolism. We provide the first direct evidence that these indole pigments form spontaneously from indolepyruvate and tryptophan without any enzymatic activity. This suggests that compounds with a proposed function in M. furfur-associated disease consist of indolepyruvate-derived spontaneously generated metabolic by-products.

J Agric Food Chem. 2008 Mar 12; 56(5): 1611-8
Andersen CM, Andersen LT, Hansen AM, Skibsted LH, Petersen MA

Degradation of the potential photosensitizers, riboflavin, chlorophyll, and porphyrin, in Danbo cheese by monochromatic light of wavelength 366, 436, or 546 nm was studied. Three cheeses were investigated, two conventional (16% fat and 25% fat) and one "organic" (25% fat). The effect of illumination was measured by fluorescence spectroscopy and analyzed using multiway and multivariate data analysis. Riboflavin was found to degrade only by 436 nm light, whereas chlorophylls and porphyrins also were influenced by 436 and 546 nm light. The organic cheese had the largest chlorophyll content both before and after similar light exposure, and no change in chlorophyll of this cheese was observed for any of the illumination wavelengths. Upon light exposure of the cheeses, volatile compounds were formed, as analyzed by gas chromatography-mass spectrometry (GC-MS). The relative concentrations of methyl Butanoate, 1-pentanol, benzaldehyde, 2-butanone, 2-heptanone, and butyl acetate were found to weakly correlate with the surface fluorescence intensity. 1-Pentanol and the ketones are secondary lipid oxidation products, consistent with a chemical coupling between photosensitizer degradation and formation of volatile lipid oxidation products.

Oncol Rep. 2008 Jan; 19(1): 245-51
Gmeiner WH, Hellmann GM, Shen P

The goal of this study was to identify systematic alterations in key cell signaling and metabolic pathways that occur during colon cancer carcinogenesis and metastasis. Understanding gene expression changes in the context of specific pathways may increase our understanding of carcinogenesis and help guide treatment. Ten cases, with matched controls, were profiled for expression of >18,000 human transcripts using Affymetrix U133A chips. Data were filtered using GeneSifter. Gene expression levels for primary colon samples were compared to a normal colon while metastatic tissues were compared to the primary colon. Differentially regulated genes were associated using the Kyoto encyclopedia of genes and genome pathways to identify cell signaling and metabolic pathways altered during carcinogenesis and metastasis. Primary colon samples displayed high positive z-scores (indicating a gene ontology term that occurs more frequently than expected) for genes involved in Wnt-signaling (4.11), nitrogen metabolism (7.30) and inositol phosphate metabolism (2.47). Expression level changes for individual genes in each cluster were statistically significant (e.g. p=0.017 for cyclin D1 in the Wnt-signaling cluster). Metastatic tissue from the liver and omentum, but not the lung, displayed a decreased expression of genes important for oxidative phosphorylation. The metastatic tissue from all sites displayed a substantially decreased expression for genes involved in Butanoate and propanoate metabolism and valine, leucine and isoleucine degradation. Our results demonstrate that systematic changes in gene expression occur for proteins involved in key cell signaling and metabolic pathways during the course of carcinogenesis and metastasis. These expression level changes complement the spectrum of mutations that characterize the progression of colorectal cancer.

Molecules. 2006; 11(1): 72-80
Sapijanskaite B, Mickevicius Y, Mikulskiene G

The reactions in benzene of 9-alkyl-3-aminocarbazoles with ethyl-3-oxoButanoate yielded ethyl-3-[(9-alkyl-9H-carbazol-3-yl)amino]but-2-enoate condensation products or N-(9-ethyl-9H-carbazol-3-yl)-3-oxobutanamide acylation products. The condensation products were cyclized to the corresponding 4,7-dihydro-pyrido[2,3-c]-carbazol-1-ones upon heating in mineral oil at 240-250 degrees C. The structures of the synthesized compounds were investigated by IR, mass spectrometry, (1)H- and (13)C-NMR spectroscopy and MM2 molecular mechanics and AM1 semi-empirical quantum mechanical methods.

Food Addit Contam. 2007 Nov; 24(11): 1306-17
Ducruet V, Vitrac O, Saillard P, Guichard E, Feigenbaum A, Fournier N

The sorption of 14 aroma compounds into PET and PVC was monitored during storage of a strawberry syrup for 1 year. Concentrations in the syrup and in the polymer were determined during storage and compared with previously published results obtained with glass bottles. Apparent partition coefficients between the polymer and the syrup (noted K app) were estimated from experimental kinetics without reaching equilibrium K app values and optimally identified from the kinetic data obtained between 30 and 90 days. They exhibited a similar behaviour for both polymers with values were between 2 x 10(-5) and 2 x 10(-3), 4 x 10(-5) and 3 x 10(-2), respectively, for PET and PVC. The variation of K app values in PET was mainly correlated to the polarity of tested compounds as assessed by their log P values. By contrast, the variations in K app values for PVC were mainly related to their chain lengths. Due to slightly higher partition coefficients and diffusion coefficients in PVC compared with PET, the amount of absorbed aroma was four times higher in PVC than in PET; however, the amount of absorbed aroma compounds was less than 0.1% of the initial amount present into the syrup, except for octyl Butanoate. The variation in concentration in the syrup was interpreted as a combination of a degradation process and a transport process into the packaging material. Both effects were particularly noticeable for both PET and unstable aroma compounds.

Bioorg Med Chem. 2007 Sep 15; 15(18): 6037-42
Wimmer Z, Floro AJ, Zarevúcka M, Wimmerová M, Sello G, Orsini F

During the investigation of ester derivatives (juvenogens, biochemically activated insect hormonogenic compounds) of biologically active alcohols with potential application in insect pest control, a need for availability of all existing stereoisomers of ethyl N-{2-[4-(2-butanoyloxycyclohexyl)methyl]phenoxy}ethyl carbamate occurred. They were synthesized from their chiral precursors, the corresponding stereoisomers of 2-(4-methoxybenzyl)cyclohexyl Butanoate, by removing their protecting group (methyl), and by subsequent condensation of the aromatic hydroxyl moiety with ethyl N-(2-bromoethyl) carbamate. The requested enantiomers of 2-(4-methoxybenzyl)cyclohexyl Butanoate were obtained by a Candida antarctica lipase-mediated transesterification and chiral resolution of the respective racemic cis- and trans-isomers of 2-(4-methoxybenzyl)cyclohexanol either directly or after a subsequent chemical esterification of the chiral precursor. In this synthesis, two convenient butanoic acid activating esters, vinyl Butanoate and 2,2,2-trifluoroethyl Butanoate, were employed, and the chiral precursors in the synthesis of the target molecules were obtained in 41-48% yields (i.e., 82-96% conversion), and with enantiomeric purity ee=96-98%, respectively. The enantiomeric purity of the products was determined by chiral HPLC analysis, and their absolute configuration was assigned on the basis of analyzing the (1)H and (19)F NMR spectra of their diastereoisomeric Mosher acid (3,3,3-trifluoromethyl-2-methoxy-2-phenylpropanoic acid) esters.

Org Biomol Chem. 2007 May 21; 5(10): 1615-20
Komoda T, Sugiyama Y, Hirota A

The biosynthesis of tetrapetalones (tetrapetalones A, B, C, and D) in Streptomyces sp. USF-4727 was studied by feeding experiments with [1-13C] sodium propanoate, [1-13C] sodium Butanoate, [carbonyl-13C] 3-amino-5-hydroxybenzoic acid (AHBA) hydrochloride, and [1-13C] glucose, followed by analysis of the 13C-NMR spectra. These feeding experiments revealed that the four tetrapetalones were polyketide compounds constructed from propanoate, Butanoate, AHBA, and glucose. The tetrapetalone biosynthetic pathway was also suggested in this study. In this pathway, tetrapetalone A (1) is synthesized by polyketide synthase (PKS) using AHBA as a starter unit, then the side chain of 1 is subjected to acetoxylation to produce tetrapetalone B (2). Additionally, 1 is oxidized and transformed into tetrapetalone C (3). In a similar way, 2 is converted to tetrapetalone D (4). Therefore, the biosynthetic relationship of the four tetrapetalones was indicated.

Appl Microbiol Biotechnol. 2007 Jul; 75(6): 1249-56
Itoh N, Nakamura M, Inoue K, Makino Y

An asymmetric hydrogen-transfer biocatalyst consisting of mutated Rhodococcus phenylacetaldehyde reductase (PAR) or Leifsonia alcohol dehydrogenase (LSADH) was applied for some water-soluble ketone substrates. Among them, 4-hydroxy-2-butanone was reduced to (S)/(R)-1,3-butanediol, a useful intermediate for pharmaceuticals, with a high yield and stereoselectivity. Intact Escherichia coli cells overexpressing mutated PAR (Sar268) or LSADH were directly immobilized with polyethyleneimine or 1,6-diaminehexane and glutaraldehyde and evaluated in a batch reaction. This system produced (S)-1,3-butanediol [87% enantiomeric excess (e.e.)] with a space time yield (STY) of 12.5 mg h(-1) ml(-1) catalyst or (R)-1,3-butanediol (99% e.e.) with an STY of 60.3 mg h(-1) ml(-1) catalyst, respectively. The immobilized cells in a packed bed reactor continuously produced (R)-1,3-butanediol with a yield of 99% (about 49.5 g/l) from 5% (w/v) 4-hydroxy-2-Butanoate over 500 h.

Protein Pept Lett. 2007; 14(2): 165-9
Farias T, Mandrich L, Rossi M, Manco G

Previously we characterized an acetyl-esterase from Escherichia coli, formally Aes, from a thermodynamic point of view in comparative studies with thermophilic homologs. Since the enzyme appeared unusually resistant to the thermal denaturation we analysed the kinetic behaviour with respect to the temperature. The enzyme displays a surprising optimal temperature at 65 degrees C, showing a specific activity of 250 U/mg using pNP-Butanoate as substrate, but a low kinetic stability at the same temperature (t(1/2) of inactivation=5 min). By a random mutagenesis approach we searched for mutated versions of Aes with increased thermostability. We found the mutant T74A, which shows the same specific activity of wild type but a t(1/2) of inactivation of 30 min at 65 degrees C.