(Below N is a link to NCBI taxonomic web page and E link to ESTHER at designed phylum.) > cellular organisms: NE > Bacteria: NE > Terrabacteria group: NE > Actinobacteria [phylum]: NE > Actinobacteria [class]: NE > Streptomycetales: NE > Streptomycetaceae: NE > Streptomyces: NE > Streptomyces cinnamoneus group: NE > Streptomyces cinnamoneus: NE
LegendThis sequence has been compared to family alignement (MSA) red => minority aminoacid blue => majority aminoacid color intensity => conservation rate title => sequence position(MSA position)aminoacid rate Catalytic site Catalytic site in the MSA MIMISSMRKTSRRSFVAAALLAAFAAPQVPAAAAAAPAAAPATAGQEVPA PATRIPLGTKTLHLVDASRQDPWKPSAGNRELMVTLWYPSLPSREPAAPY VSKPLSRAVLGNDVLAGVRTHAVAGARPAPVPRPLVVLSPGFGMSRITLT ALGEDLASRGYAVAAVDHTYEAPVEFPGGRIEKCTLCDDSRMDPGAVVRN RAKDLRFVLDRLTGPGSELRVDARRIGVAGHSIGGASAVEVMREDRRVDA AINLDGNFFTEPPAEGLNKPVLLLGARRSGLPEPQENWERAWKQLTGWKR WLDVPAGGHMTFTDVPWIVDRFGMPGQIPPEQVEGQLGTVSAARATAVTR NYVAAFFDRHLRGRPSPLLDRPSSAHPEVTFMK
References
Title: Enhancing the secretion of a feruloyl esterase in Bacillus subtilis by signal peptide screening and rational design Liu P, Guo J, Miao L, Liu H Ref: Protein Expr Purif, :106165, 2022 : PubMed
Feruloyl esterase is a subclass of alpha/beta hydrolase, which could release ferulic acid from biomass residues for use as an efficient additive in food or pharmaceutical industries. In the present study, a feruloyl esterase with broad substrate specificity was characterised and secreted by Bacillus subtilis WB600. After codon usage optimisation and signal peptide library screening, the secretion amount of feruloyl esterase was enhanced by up to 10.2-fold in comparison with the base strain. The site-specific amino acid substitutions that facilitate protein folding further improved the secretion by about 1.5-fold. The purified rationally designed enzyme exhibited maximal activity against methyl ferulate at pH 6.5 and 65 degreesC. In the solid-state fermentation, the genetically engineered B. subtilis released about 37% of the total alkali-extractable ferulic acid in maize bran. This study provides a promising candidate for ferulic acid production and demonstrates that the secretion of a heterologous enzyme from B. subtilis can be cumulatively improved by changes in protein sequence features.
Title: Loop of Streptomyces Feruloyl Esterase Plays an Important Role in the Enzyme's Catalyzing the Release of Ferulic Acid from Biomass Uraji M, Tamura H, Mizohata E, Arima J, Wan K, Ogawa K, Inoue T, Hatanaka T Ref: Applied Environmental Microbiology, 84:, 2018 : PubMed
Feruloyl esterases (FAEs) are key enzymes required for the production of ferulic acid from agricultural biomass. Previously, we identified and characterized R18, an FAE from Streptomyces cinnamoneus NBRC 12852, which showed no sequence similarity to the known FAEs. To determine the region involved in its catalytic activity, we constructed chimeric enzymes using R18 and its homolog (TH2-18) from S. cinnamoneus strain TH-2. Although R18 and TH2-18 showed 74% identity in their primary sequences, the recombinant proteins of these two FAEs (recombinant R18 [rR18] and rTH2-18) showed very different specific activities toward ethyl ferulate. By comparing the catalytic activities of the chimeras, a domain comprised of residues 140 to 154 was found to be crucial for the catalytic activity of R18. Furthermore, we analyzed the crystal structure of rR18 at a resolution of 1.5 A to elucidate the relationship between its activity and its structure. rR18 possessed a typical catalytic triad, consisting of Ser-191, Asp-214, and His-268, which was characteristic of the serine esterase family. By structural analysis, the above-described domain was found to be present in a loop-like structure (the R18 loop), which possessed a disulfide bond conserved in the genus Streptomyces Moreover, compared to rTH2-18 of its parental strain, the TH2-18 mutant, in which Pro and Gly residues were inserted into the domain responsible for forming the R18 loop, showed markedly high kcat values using artificial substrates. We also showed that the FAE activity of TH2-18 toward corn bran, a natural substrate, was improved by the insertion of the Gly and Pro residues.IMPORTANCEStreptomyces species are widely distributed bacteria that are predominantly present in soil and function as decomposers in natural environments. They produce various enzymes, such as carbohydrate hydrolases, esterases, and peptidases, which decompose agricultural biomass. In this study, based on the genetic information on two Streptomyces cinnamoneus strains, we identified novel feruloyl esterases (FAEs) capable of producing ferulic acid from biomass. These two FAEs shared high similarity in their amino acid sequences but did not resemblance any known FAEs. By comparing chimeric proteins and performing crystal structure analysis, we confirmed that a flexible loop was important for the catalytic activity of Streptomyces FAEs. Furthermore, we determined that the catalytic activity of one FAE was improved drastically by inserting only 2 amino acids into its loop-forming domain. Thus, differences in the amino acid sequence of the loop resulted in different catalytic activities. In conclusion, our findings provide a foundation for the development of novel enzymes for industrial use.
Title: Application of two newly identified and characterized feruloyl esterases from Streptomyces sp. in the enzymatic production of ferulic acid from agricultural biomass Uraji M, Arima J, Inoue Y, Harazono K, Hatanaka T Ref: PLoS ONE, 9:e104584, 2014 : PubMed
Ferulic acid (FA), a component of hemicellulose in plant cell walls, is a phenolic acid with several potential applications based on its antioxidant properties. Recent studies have shown that feruloyl esterase (FAE) is a key bacterial enzyme involved in FA production from agricultural biomass. In this study, we screened a library of 43 esterases from Streptomyces species and identified two enzymes, R18 and R43, that have FAE activity toward ethyl ferulate. In addition, we characterized their enzyme properties in detail. R18 and R43 showed esterase activity toward other hydroxycinnamic acid esters as well, such as methyl p-coumarate, methyl caffeate, and methyl sinapinate. The amino acid sequences of R18 and R43 were neither similar to each other, nor to other FAEs. We found that R18 and R43 individually showed the ability to produce FA from corn bran; however, combination with other Streptomyces enzymes, namely xylanase and alpha-l-arabinofuranosidase, increased FA production from biomass such as corn bran, defatted rice bran, and wheat bran. These results suggest that R18 and R43 are effective FAEs for the enzymatic production of FA from biomass.
Streptomyces cinnamoneus (Streptoverticillium cinnamoneum) estA, R18 Feruoyl esterase strain: NBRC 12852 R18
