Feruloyl esterase immobilization in mesoporous silica particles and characterization in hydrolysis and transesterification

Sethumadhavan

et al. 2021

Sci Rep

Plant polyphenol gossypol has anticancer activities. This may increase cottonseed value by using gossypol as a health intervention agent. It is necessary to understand its molecular mechanisms before human consumption. The aim was to uncover the effects of gossypol on cell viability and gene expression in cancer cells. In this study, human colon cancer cells (COLO 225) were treated with gossypol. MTT assay showed significant inhibitory effect under high concentration and longtime treatment. We analyzed the expression of 55 genes at the mRNA level in the cells; many of them are regulated by gossypol or ZFP36/TTP in cancer cells. BCL2 mRNA was the most stable among the 55 mRNAs analyzed in human colon cancer cells. GAPDH and RPL32 mRNAs were not good qPCR references for the colon cancer cells. Gossypol decreased the mRNA levels of DGAT, GLUT, TTP, IL families and a number of previously reported genes. In particular, gossypol suppressed the expression of genes coding for CLAUDIN1, ELK1, FAS, GAPDH, IL2, IL8 and ZFAND5 mRNAs, but enhanced the expression of the gene coding for GLUT3 mRNA. The results showed that gossypol inhibited cell survival with decreased expression of a number of genes in the colon cancer cells.

Section: Discussionmentioning

confidence: 90%

“…DGATs are divided into DGAT1 and DGAT2 subfamilies in animals and DGAT3 subfamily are present in plants 52 – 55 . DGAT2 mRNA was the major form of DGAT mRNAs in the mouse adipocytes and macrophages 56 , 57 . Gossypol was shown to be a strong stimulator of DGAT2 gene expression in mouse macrophages 56 .…”

Section: Resultsmentioning

confidence: 98%

Gossypol decreased cell viability and down-regulated the expression of a number of genes in human colon cancer cells

Sethumadhavan

et al. 2021

Sci Rep

“…The immobilization support used in this study was Santa Barbara Amorphous type 15 (SBA-15), a type of mesoporous silica (MPS) particles, which present a tunable and ordered network of hexagonal porous structures (Zhao et al 1998). They were synthesized and characterized as previously described (Bonzom et al 2018), and were a kind gift from Milene Zezzi Do Valle Gomes (Chalmers University of Technology). The main properties of the particles are: pore size 10.1 nm, BET surface area 439 m 2 /g and specific pore volume 1.11 cm 3 /g.…”

Section: Methodsmentioning

confidence: 99%

Glycosylation influences activity, stability and immobilization of the feruloyl esterase 1a from Myceliophthora thermophila

et al. 2019

Self Cite

Heterologous protein production is widely used in industrial biotechnology. However, using non-native production hosts can lead to enzymes with altered post-translational modifications, such as glycosylation. We have investigated how production in a non-native host affects the physicochemical properties and enzymatic activity of a feruloyl esterase from Myceliophthora thermophila , Mt Fae1a. The enzyme was produced in two microorganisms that introduce glycosylation ( M. thermophila and Pichia pastoris ) and in Escherichia coli (non-glycosylated). Mass spectrometric analysis confirmed the presence of glycosylation and revealed differences in the lengths of glycan chains between the enzymes produced in M. thermophila and P. pastoris . The melting temperature and the optimal temperature for activity of the non-glycosylated enzyme were considerably lower than those of the glycosylated enzymes. The three Mt Fae1a versions also exhibited differences in specific activity and specificity. The catalytic efficiency of the glycosylated enzymes were more than 10 times higher than that of the non-glycosylated one. In biotechnology, immobilization is often used to allow reusing enzyme and was investigated on mesoporous silica particles. We found the binding kinetics and immobilization yield differed between the enzyme versions. The largest differences were observed when comparing enzymes with and without glycosylation, but significant variations were also observed between the two differently glycosylated enzymes. We conclude that the biotechnological value of an enzyme can be optimized for a specific application by carefully selecting the production host. Electronic supplementary material The online version of this article (10.1186/s13568-019-0852-z) contains supplementary material, which is available to authorized users.

“…[27], which had 40-fold higher K M values and around 60-fold lower k cat values. Compared to the fungal FAE AnFaeA, from Aspergillus niger, and the commercial enzyme E-FAERU, from a rumen microorganism, FjCE1 possessed the smallest K M value at 0.11 mM, compared to 0.78 and 0.43 mM for AnFaeA and E-FAERU, respectively [28,29]. However, both AnFaeA and E-FAERU possessed higher k cat values (70.74 and 31.4 s −1 , respectively), resulting in three to fourfold better catalytic efficiencies than those observed for FjCE1.…”

Section: Biochemical Characterization Of the F Johnsoniae Ce6-ce1 Enmentioning

confidence: 99%

Multimodular fused acetyl–feruloyl esterases from soil and gut Bacteroidetes improve xylanase depolymerization of recalcitrant biomass

Kmezik

Bonzom

Olsson

et al. 2020

Biotechnol Biofuels

Self Cite

Background: Plant biomass is an abundant and renewable carbon source that is recalcitrant towards both chemical and biochemical degradation. Xylan is the second most abundant polysaccharide in biomass after cellulose, and it possesses a variety of carbohydrate substitutions and non-carbohydrate decorations which can impede enzymatic degradation by glycoside hydrolases. Carbohydrate esterases are able to cleave the ester-linked decorations and thereby improve the accessibility of the xylan backbone to glycoside hydrolases, thus improving the degradation process. Enzymes comprising multiple catalytic glycoside hydrolase domains on the same polypeptide have previously been shown to exhibit intramolecular synergism during degradation of biomass. Similarly, natively fused carbohydrate esterase domains are encoded by certain bacteria, but whether these enzymes can result in similar synergistic boosts in biomass degradation has not previously been evaluated. Results: Two carbohydrate esterases with similar architectures, each comprising two distinct physically linked catalytic domains from families 1 (CE1) and 6 (CE6), were selected from xylan-targeting polysaccharide utilization loci (PULs) encoded by the Bacteroidetes species Bacteroides ovatus and Flavobacterium johnsoniae. The full-length enzymes as well as the individual catalytic domains showed activity on a range of synthetic model substrates, corn cob biomass, and Japanese beechwood biomass, with predominant acetyl esterase activity for the N-terminal CE6 domains and feruloyl esterase activity for the C-terminal CE1 domains. Moreover, several of the enzyme constructs were able to substantially boost the performance of a commercially available xylanase on corn cob biomass (close to twofold) and Japanese beechwood biomass (up to 20-fold). Interestingly, a significant improvement in xylanase biomass degradation was observed following addition of the full-length multidomain enzyme from B. ovatus versus the addition of its two separated single domains, indicating an intramolecular synergy between the esterase domains. Despite high sequence similarities between the esterase domains from B. ovatus and F. johnsoniae, their addition to the xylanolytic reaction led to different degradation patterns. Conclusion: We demonstrated that multidomain carbohydrate esterases, targeting the non-carbohydrate decorations on different xylan polysaccharides, can considerably facilitate glycoside hydrolase-mediated hydrolysis of xylan