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|Substrate For (Enzyme):
|Spectrophotometer (Semi-quantitative), Petri-dish (Qualitative)
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High purity dyed and crosslinked insoluble AZCL-Galactomannan (Carob) for identification of enzyme activities in research, microbiological enzyme assays and in vitro diagnostic analysis.
Substrate for the assay of endo-1,4-β-D-mannanase.
See our other AZCL-dyed substrates.
McCleary, B. V. (1978). Carbohydrate Research, 67(1), 213-221.
A simple assay procedure for β-D-mannanase enzyme has been developed which employs carob D-galacto-D-mannan dyed with Remazolbrilliant Blue. Additionally, the procedure is quantitative, relatively sensitive, and highly specific for β-D-mannanase enzyme. It can be readily used for the determination of β-D-mannanase activity in crude enzyme preparations and column-chromatography eluates.Hide Abstract
Non-digestible galactomannan oligosaccharides from Cassia seed gum modulate microbiota composition and metabolites of human fecal inoculum.
Miao, M., Shi, Y., Li, Y., Jiang, Z., Liu, J. & Yang, S. (2021). Journal of Functional Foods, 86, 104705.
The digestibility and prebiotic potential of galactomannan oligosaccharides prepared from Cassia seed gum (CMOS) was studied. CMOS were tolerant against simulated gastrointestinal digestion. During in vitro human fecal fermentation with CMOS, the microbiota composition was significantly changed with promoted growth of potential beneficial genera (i.e. Bifidobacterium, Lactobacillus, Bacteroides, Veillonella) while that of potential harmful genera (i.e. Fusobacterium, Lachnospiraceae, and Sutterella) was inhibited. Short chain fatty acids were produced with decreased medium pH, while acetic acid (20.85 mM) and propionic acid (19.77 mM) were the predominant metabolites. For CMOS utilization, α-galactosidase and β-mannosidase were involved with the enzyme activity reached the maximum after 3 h fermentation. The HPAEC chromatograms showed that CMOS with DP = 3 and 4 were prior utilized through enzymatic hydrolysis within 36 h fermentation. Findings gained here will introduce the utilization and targeted production of CMOS (DP = 3 and 4) as prebiotics with defined structural features.Hide Abstract
Pitt, J. I., Lange, L., Lacey, A. E., Vuong, D., Midgley, D. J., Greenfield, P., Bradbury, M. I., Lacey, E., Busk, P. K., Pilgaard, B., Chooi, Y. H. & Piggott, A. M. (2017). PloS One, 12(4), e0170254.
Aspergillus hancockii sp. nov., classified in Aspergillus subgenus Circumdati section Flavi, was originally isolated from soil in peanut fields near Kumbia, in the South Burnett region of southeast Queensland, Australia, and has since been found occasionally from other substrates and locations in southeast Australia. It is phylogenetically and phenotypically related most closely to A. leporis States and M. Chr., but differs in conidial colour, other minor features and particularly in metabolite profile. When cultivated on rice as an optimal substrate, A. hancockii produced an extensive array of 69 secondary metabolites. Eleven of the 15 most abundant secondary metabolites, constituting 90% of the total area under the curve of the HPLC trace of the crude extract, were novel. The genome of A. hancockii, approximately 40 Mbp, was sequenced and mined for genes encoding carbohydrate degrading enzymes identified the presence of more than 370 genes in 114 gene clusters, demonstrating that A. hancockii has the capacity to degrade cellulose, hemicellulose, lignin, pectin, starch, chitin, cutin and fructan as nutrient sources. Like most Aspergillus species, A. hancockii exhibited a diverse secondary metabolite gene profile, encoding 26 polyketide synthase, 16 nonribosomal peptide synthase and 15 nonribosomal peptide synthase-like enzymes.Hide Abstract
Prakash, R., Hallett, I. C., Wong, S. F., Johnston, S. L., O’Donoghue, E. M., McAtee, P. A., Seal, A. G., Atkinson, R. G. & Schröder, R. (2017). BMC Plant Biology, 17(1), 86.
Background: Unlike in abscission or dehiscence, fruit of kiwifruit Actinidia eriantha develop the ability for peel detachment when they are ripe and soft in the absence of a morphologically identifiable abscission zone. Two closely-related genotypes with contrasting detachment behaviour have been identified. The ‘good-peeling’ genotype has detachment with clean debonding of cells, and a peel tissue that does not tear. The ‘poor-peeling’ genotype has poor detachability, with cells that rupture upon debonding, and peel tissue that fragments easily. Results: Structural studies indicated that peel detachability in both genotypes occurred in the outer pericarp beneath the hypodermis. Immunolabelling showed differences in methylesterification of pectin, where the interface of labelling coincided with the location of detachment in the good-peeling genotype, whereas in the poor-peeling genotype, no such interface existed. This zone of difference in methylesterification was enhanced by differential cell wall changes between the peel and outer pericarp tissue. Although both genotypes expressed two polygalacturonase genes, no enzyme activity was detected in the good-peeling genotype, suggesting limited pectin breakdown, keeping cell walls strong without tearing or fragmentation of the peel and flesh upon detachment. Differences in location and amounts of wall-stiffening galactan in the peel of the good-peeling genotype possibly contributed to this phenotype. Hemicellulose-acting transglycosylases were more active in the good-peeling genotype, suggesting an influence on peel flexibility by remodelling their substrates during development of detachability. High xyloglucanase activity in the peel of the good-peeling genotype may contribute by having a strengthening effect on the cellulose-xyloglucan network. Conclusions: In fruit of A. eriantha, peel detachability is due to the establishment of a zone of discontinuity created by differential cell wall changes in peel and outer pericarp tissues that lead to changes in mechanical properties of the peel. During ripening, the peel becomes flexible and the cells continue to adhere strongly to each other, preventing breakage, whereas the underlying outer pericarp loses cell wall strength as softening proceeds. Together these results reveal a novel and interesting mechanism for enabling cell separation.Hide Abstract
Huang, Y., Yi, Z., Jin, Y., Huang, M., He, K., Liu, D., Luo, H., Zhao, D., He, H., Fang, Y. & Zhao, H. (2017). Frontiers in Microbiology, 8, 1747.
Chinese liquor is one of the world's best-known distilled spirits and is the largest spirit category by sales. The unique and traditional solid-state fermentation technology used to produce Chinese liquor has been in continuous use for several thousand years. The diverse and dynamic microbial community in a liquor starter is the main contributor to liquor brewing. However, little is known about the ecological distribution and functional importance of these community members. In this study, metatranscriptomics was used to comprehensively explore the active microbial community members and key transcripts with significant functions in the liquor starter production process. Fungi were found to be the most abundant and active community members. A total of 932 carbohydrate-active enzymes, including highly expressed auxiliary activity family 9 and 10 proteins, were identified at 62°C under aerobic conditions. Some potential thermostable enzymes were identified at 50, 62, and 25°C (mature stage). Increased content and overexpressed key enzymes involved in glycolysis and starch, pyruvate and ethanol metabolism were detected at 50 and 62°C. The key enzymes of the citrate cycle were up-regulated at 62°C, and their abundant derivatives are crucial for flavor generation. Here, the metabolism and functional enzymes of the active microbial communities in NF liquor starter were studied, which could pave the way to initiate improvements in liquor quality and to discover microbes that produce novel enzymes or high-value added products.Hide Abstract
Carrillo-Barral, N., Matilla, A. J., del Carmen Rodríguez-Gacio, M. & Iglesias-Fernández, R. (2017). Planta, 1-13.
The contribution of endo-β-mannanase (MAN) genes to the germination of the wild-type Sisymbrium officinale and cultivated Brassica rapa (Brassicaceae) species has been explored. In both species, mannans have been localized to the imbibed external seed coat layer (mucilage) by fluorescence immunolocalization and MAN enzymatic activity increases in seeds as imbibition progresses, reaching a peak before 100% germination is achieved. The MAN gene families have been annotated and the expression of their members analyzed in vegetative and reproductive organs. In S. officinale and B. rapa, MAN2, MAN5, MAN6, and MAN7 transcripts accumulate upon seed imbibition. SoMAN7 is the most expressed MAN gene in S. officinale germinating seeds, as occurs with its ortholog in Arabidopsis thaliana, but in B. rapa, the most abundant transcripts are BrMAN2 and BrMAN5. These genes (MAN2, MAN5, MAN6, and MAN7) are localized, by mRNA in situ hybridization, to the micropylar at the endosperm layer and to the radicle in S. officinale, but in B. rapa, these mRNAs are faintly found to the micropylar living seed coat layer and are mainly present at the radicle tip and the vascular bundles. If the domestication process undergone by B. rapa is responsible for these different MAN expression patterns, upon germination remains to be elucidated. Since mannans and MAN genes are not spatially distributed in the same seed tissues, a movement of MAN enzymes that are synthesized with typical signal peptides from the embryo tissues to the mucilage layer (via apoplastic space) is necessary for the mannans to be hydrolyzed.Hide Abstract
Zhao, Y., Song, D., Sun, J. & Li, L. (2013). The Plant Journal, 74(3), 473-485.
Endo-1,4-β-mannanase is known to able to hydrolyze mannan-type polysaccharides in cell wall remodeling, but its function in regulating wall thickening has been little studied. Here we show that a Populus endo-1,4-β-mannanase gene, named PtrMAN6, suppresses cell wall thickening during xylem differentiation. PtrMAN6 is expressed specifically in xylem tissue and its encoded protein localizes to developing vessel cells. Overexpression of PtrMAN6 enhanced wall loosening as well as suppressed secondary wall thickening, whilst knockdown of its expression promoted secondary wall thickening. Transcriptional analysis revealed that PtrMAN6 overexpression downregulated the transcriptional program of secondary cell wall thickening, whilst PtrMAN6 knockdown upregulated transcriptional activities toward secondary wall formation. Activity of PtrMAN6 hydrolysis resulted in the generation of oligosaccharide compounds from cell wall polysaccharides. Application of the oligosaccharides resulted in cellular and transcriptional changes that were similar to those found in PtrMAN6 overexpressed transgenic plants. Overall, our results demonstrated that PtrMAN6 plays a role in hydrolysis of mannan-type wall polysaccharides to produce oligosaccharides that may serve as signaling molecules to suppress cell wall thickening during wood xylem cell differentiation.Hide Abstract
Lund, M. B., Schätzle, S., Schramm, A. & Kjeldsen, K. U. (2012). Antonie van Leeuwenhoek, 101(3), 507-514.
Clone library-based studies have shown that almost all lumbricid earthworm species harbour host-specific symbiotic bacteria belonging to the novel genus Verminephrobacter in their nephridia (excretory organs). To date the only described representative from this genus is Verminephrobacter eiseniae, the specific symbiont of the earthworm Eisenia fetida. In this study two novel rod-shaped, non-endosporeforming, betaproteobacterial symbionts were isolated from the nephridia of two closely related earthworm species. Both isolates were affiliated with the genus Verminephrobacter by 16SrRNA gene sequence analysis. Similarly to V. eiseniae, the two isolates grew aerobically with a preference for low oxygen concentrations on a range of sugars, fatty acids and amino acids and fermentatively on glucose and pyruvate. These phenotypes match well with the conditions reported or inferred for the nephridial environment. Based on 16S rRNA gene similarity, DNA–DNA hybridization value and phenotypic characteristics the two isolates are clearly distinct from V. eiseniae. Phenotypic characteristics could not clearly differentiate the two strains as separate species but a low DNA–DNA hybridization value of 57.3%, their earthworm host specificity, differing temperature ranges and pH optima suggest that they represent two subspecies of a novel species of Verminephrobacter. For this species, the name V. aporrectodeae sp. nov. is proposed, with the two subspecies V. aporrectodeae subsp. tuberculatae (type strain, At4T = DSM 21361T = LMG 25313T) and V. aporrectodeae subsp. caliginosae (type strain, Ac9T = DSM 21895T = LMG 25312T) isolated from the nephridia of the earthworms Aporrectodea tuberculate and A. caliginosa, respectively.Hide Abstract
Schröder, R., Wegrzyn, T. F., Sharma, N. N. & Atkinson, R. G. (2006). Planta, 224(5), 1091-1102.
Mannan transglycosylases are cell wall enzymes able to transfer part of the mannan polysaccharide backbone to mannan-derived oligosaccharides (Schröder et al. in Planta 219:590–600, 2004). Mannan transglycosylase activity was purified to near homogeneity from ripe tomato fruit. N-terminal sequencing showed that the dominant band seen on SDS-PAGE was identical to LeMAN4a, a hydrolytic endo-β-mannanase found in ripe tomato fruit (Bewley et al. in J Exp Bot 51:529–538, 2000). Recombinant LeMAN4a protein expressed in Escherichia coli exhibited both mannan hydrolase and mannan transglycosylase activity. Western analysis of ripe tomato fruit tissue using an antibody raised against tomato seed endo-β-mannanase revealed four isoforms present after 2D-gel electrophoresis in the pH range 6–11. On separation by preparative liquid isoelectric focussing, these native isoforms exhibited different preferences for transglycosylation and hydrolysis. These results demonstrate that endo-β-mannanase has two activities: it can either hydrolyse mannan polysaccharides, or in the presence of mannan-derived oligosaccharides, carry out a transglycosylation reaction. We therefore propose that endo-β-mannanase should be renamed mannan transglycosylase/hydrolase, in accordance with the nomenclature established for xyloglucan endotransglucosylase/hydrolase. The role of endo-acting mannanases in modifying the structure of plant cell walls during cell expansion, seed germination and fruit ripening may need to be reinterpreted in light of their potential action as transglycosylating or hydrolysing enzymes.Hide Abstract
Misko, A. L. & Germida, J. J. (2002). FEMS Microbiology Ecology, 42(3), 399-407.
Among the most important rhizosphere bacteria are the pseudomonads, which are aggressive colonizers and utilize a wide range of substrates as carbon sources. The objective of this study was to determine if the taxonomic or metabolic diversity of pseudomonads differed among field-grown canola cultivars. Bacteria (n=2257) were isolated from the rhizosphere and root interior of six cultivars of field-grown canola, including three transgenic varieties. The bacteria were identified by fatty acid methyl ester (FAME) analysis, and about 35% were identified as Pseudomonas species. The most abundant species were Pseudomonas putida and Pseudomonas chlororaphis. Dendrograms based on FAME analysis revealed that many pseudomonad strains were found in all of the canola cultivars. Pseudomonads of the same strain were found in both the rhizosphere and the root interior of canola plants, suggesting that endophytic bacteria were a subset of the rhizosphere community. Because metabolic profiling provides more useful information than taxonomy, P. putida and P. chlororaphis isolates were characterized for their ability to utilize carbon substrates and produce several enzymes. Bacteria isolated from different plant cultivars had different carbon utilization profiles, but when only carbon substrates found in root exudates were analyzed, the cultivar effect was less pronounced. These characterizations also demonstrated that bacteria that were determined by FAME to be the same strain were metabolically different, suggesting functional redundancy among Pseudomonas isolates. The results of this study suggest that pseudomonads were functionally diverse. They differed in their metabolic potential among the canola cultivars from which they were isolated. Because bacteria capable of using many substrates can effectively adapt to new environments, these results have implications for the use of pseudomonads as biofertilizers, biological control agents and plant growth-promoting bacteria in canola.Hide Abstract