Identification and functional characterization of levS, a gene encoding for a levansucrase from Leuconostoc mesenteroides NRRL B-512 F

Morales-Arrieta, S.; Rodríguez, Maria E.; Segovia, Lorenzo; López-Munguı́a, Agustı́n; Olvera-Carranza, Clarita

doi:10.1016/j.gene.2006.02.007

Cited by 55 publications

(49 citation statements)

References 35 publications

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“…Although the chimeric constructions were the main and most abundant proteins, several activity bands associated with the fructan production were present. These additional activity bands of lower molecular weight than the expected chimers were the result of chimer proteolysis, a common problem associated to MD-FSs production (19,16,20). In spite of this limited proteolysis, a modification of the crude chimer specificity was observed (Fig.…”

Section: Resultsmentioning

confidence: 91%

“…Recently, a subfamily of multidomain fructansucrases (MD-FSs) that have acquired structural domains from the N-and C-terminal regions of glucansucrases (GSs) has been described in Gram-positive Leuconostoc spp. (16). These FSs include inulosucrase (IslA) from Leuconostoc citreum (19,22) and levansucrases (LevS, LevC, and LevL) from L. mesenteroides (16,20).…”

mentioning

confidence: 99%

“…(16). These FSs include inulosucrase (IslA) from Leuconostoc citreum (19,22) and levansucrases (LevS, LevC, and LevL) from L. mesenteroides (16,20).…”

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confidence: 99%

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Design of Chimeric Levansucrases with Improved Transglycosylation Activity

Olvera

Centeno-Leija

Ruiz-Leyva

et al. 2012

Appl Environ Microbiol

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Fructansucrases (FSs), including levansucrases and inulosucrases, are enzymes that synthesize fructose polymers from sucrose by the direct transfer of the fructosyl moiety to a growing polymer chain. These enzymes, particularly the single domain fructansucrases, also possess an important hydrolytic activity, which may account for as much as 70 to 80% of substrate conversion, depending on reaction conditions. Here, we report the construction of four chimeric levansucrases from SacB, a single domain levansucrase produced by Bacillus subtilis. Based on observations derived from the effect of domain deletion in both multidomain fructansucrases and glucansucrases, we attached different extensions to SacB. These extensions included the transitional domain and complete C-terminal domain of Leuconostoc citreum inulosucrase (IslA), Leuconostoc mesenteroides levansucrase (LevC), and a L. mesenteroides glucansucrase (DsrP). It was found that in some cases the hydrolytic activity was reduced to less than 10% of substrate conversion; however, all of the constructs were as stable as SacB. This shift in enzyme specificity was observed even when the SacB catalytic domain was extended only with the transitional region found in multidomain FSs. Specific kinetic analysis revealed that this change in specificity of the SacB chimeric constructs was derived from a 5-fold increase in the transfructosylation k cat and not from a reduction of the hydrolytic k cat , which remained constant.

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Section: Resultsmentioning

confidence: 91%

mentioning

confidence: 99%

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Design of Chimeric Levansucrases with Improved Transglycosylation Activity

Olvera

Centeno-Leija

Ruiz-Leyva

et al. 2012

Appl Environ Microbiol

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“…levansucrase (ec 2.4.1.10) and inulosucrase (ec 2.4.1.9) have been well described (29,31,32). Robyt and Walseth (39) reported that levansucrase is present in dextransucrase preparations obtained from Leuconostoc mesenteroides B-512F.…”

Section: Introductionmentioning

confidence: 93%

“…only strains from the Streptococcus, Lactobacillus and Leuconostoc genera are producers of both GS and FS (23,25,28,29). GS and FS from L. mesenteroides are sucrose inducible enzymes, whereas GS from Streptococcus spp.…”

Section: Introductionmentioning

confidence: 99%

Characterization of Glucansucrases and Fructansucrases Produced by Wild StrainsLeuconostoc MesenteroidesURE13 andLeuconostoc MesenteroidesLM17 Grown on Glucose or Fructose Medium as a Sole Carbon Source

Bivolarski

Vasileva

Dzhambazov

et al. 2013

Biotechnology & Biotechnological Equipment

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Purification and characterization of an intracellular levansucrase derived from Bacillus methylotrophicus SK 21.002

Zhang

Jiang

et al. 2015

Biotech and App Biochem

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An intracellular levansucrase from Bacillus methylotrophicus SK 21.002 was isolated, purified, and characterized. The final specific levansucrase activity was 135.40 U/mg protein with an 11.78-fold enrichment and a 9.28% recovery rate. The molecular weight of the enzyme was approximately 60,000 Da as evaluated by gel filtration and SDS-PAGE. Both the maximum transfructosylation and hydrolytic activities were observed at pH 6.5. The enzyme exhibited optimum transfructosylation activity at 40 °C, whereas the optimum temperature of hydrolytic activity was 45 °C. Cu(2+), Fe(2+), Zn(2+), and Ni(2+) inhibited both the transfructosylation and hydrolytic activities up to 100%, whereas Mn(2+) inhibited only hydrolytic activity. Ca(2+) and Mg(2+) stimulated both transfructosylation and hydrolytic activities. The chemical modifiers (n-bromosuccinimide and phenylmethanesulfonyl fluoride) strongly inhibited hydrolytic and transfructosylation activity of the levansucrase. The Km and Vmax values of the purified levansucrase were 117.2 mM and 33.23 μmol/mg·Min, respectively. When the fructose concentration was below 0.2 M, higher fructose concentrations promoted the transfructosylation and inhibited the hydrolytic activity.

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Identification and functional characterization of levS, a gene encoding for a levansucrase from Leuconostoc mesenteroides NRRL B-512 F

Cited by 55 publications

References 35 publications

Design of Chimeric Levansucrases with Improved Transglycosylation Activity

Design of Chimeric Levansucrases with Improved Transglycosylation Activity

Characterization of Glucansucrases and Fructansucrases Produced by Wild StrainsLeuconostoc MesenteroidesURE13 andLeuconostoc MesenteroidesLM17 Grown on Glucose or Fructose Medium as a Sole Carbon Source

Purification and characterization of an intracellular levansucrase derived from Bacillus methylotrophicus SK 21.002

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