scholarly journals Molecular-scale features that govern the effects of O-glycosylation on a carbohydrate-binding module

2015 ◽  
Vol 6 (12) ◽  
pp. 7185-7189 ◽  
Author(s):  
Xiaoyang Guan ◽  
Patrick K. Chaffey ◽  
Chen Zeng ◽  
Eric R. Greene ◽  
Liqun Chen ◽  
...  
Keyword(s):  
Amino Acid ◽  
Glycosylation Site ◽  
Glycan Structure ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Amino Acid Residues ◽  
Glycosidic Linkage ◽  
Molecular Scale

The importance of the glycan structure and size, amino acid residues near the glycosylation site, and glycosidic linkage in controlling the effects of CBMO-glycosylation is shown.

scholarly journals Novel Carbohydrate-Binding Module of β-1,3-Xylanase from a Marine Bacterium, Alcaligenes sp. Strain XY-234

2002 ◽  
Vol 184 (9) ◽  
pp. 2399-2403 ◽  
Author(s):  
Fumiyoshi Okazaki ◽  
Yutaka Tamaru ◽  
Shinnosuke Hashikawa ◽  
Yu-Teh Li ◽  
Toshiyoshi Araki
Keyword(s):  
Amino Acid ◽  
Marine Bacterium ◽  
Catalytic Domain ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Amino Acid Residues ◽  
Glycosyl Hydrolases ◽  
Binding Studies ◽  
Calculated Molecular Mass ◽  
Reading Frame

ABSTRACT A β-1,3-xylanase gene (txyA) from a marine bacterium, Alcaligenes sp. strain XY-234, has been cloned and sequenced. txyA consists of a 1,410-bp open reading frame that encodes 469 amino acid residues with a calculated molecular mass of 52,256 Da. The domain structure of the β-1,3-xylanase (TxyA) consists of a signal peptide of 22 amino acid residues, followed by a catalytic domain which belongs to family 26 of the glycosyl hydrolases, a linker region with one array of DGG and six repeats of DNGG, and a novel carbohydrate-binding module (CBM) at the C terminus. The recombinant TxyA hydrolyzed β-1,3-xylan but not other polysaccharides such as β-1,4-xylan, carboxymethylcellulose, curdlan, glucomannan, or β-1,4-mannan. TxyA was capable of binding specifically to β-1,3-xylan. The analysis using truncated TxyA lacking either the N- or C-terminal region indicated that the region encoding the CBM was located between residues 376 and 469. Binding studies on the CBM revealed that the Kd and the maximum amount of protein bound to β-1,3-xylan were 4.2 μM and 18.2 μmol/g of β-1,3-xylan, respectively. Furthermore, comparison of the enzymatic properties between proteins with and without the CBM strongly indicated that the CBM of TxyA plays an important role in the hydrolysis of β-1,3-xylan.

scholarly journals Rat mammary-gland transferrin: nucleotide sequence, phylogenetic analysis and glycan structure

1995 ◽  
Vol 307 (1) ◽  
pp. 47-55 ◽  
Author(s):  
H Escrivá ◽  
A Pierce ◽  
B Coddeville ◽  
F González ◽  
M Benaissa ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Mammary Gland ◽  
Sialic Acid ◽  
Amino Acid ◽  
Signal Sequence ◽  
Glycosylation Site ◽  
Glycan Structure ◽  
Sialic Acid Content ◽  
Native Page ◽  
Rat Mammary Gland

The complete cDNA for rat mammary-gland transferrin (Tf) has been sequenced and also the native protein isolated from milk in order to analyse the structure of the main glycan variants present. A lactating-rat mammary-gland cDNA library in lambda gt10 was screened with a partial cDNA copy of rat liver Tf and subsequently rescreened with 5′ fragments of the longest clones. This produced a 2275 bp insert coding for an open reading frame of 695 amino acid residues. This includes a 19-amino acid signal sequence and the mature protein containing 676 amino acids and one N-glycosylation site in the C-terminal domain at residue 490. Phylogenetic analysis was carried out using 14 translated Tf nucleotide sequences, and the derived evolutionary tree shows that at least three gene duplication events have occurred during Tf evolution, one of which generated the N- and C-terminal domains and occurred before separation of arthropods and chordates. The two halves of human melanotransferrin are more similar to each other than to any other sequence, which contrasts with the pattern shown by the remaining sequences. Native rat milk Tf is separated into four bands on native PAGE that differ only in their sialic acid content: one biantennary glycan is present containing either no sialic acid residues or up to three. The complete structures of the two major variants were determined by methylation, m.s. and 400 MHz 1H-n.m.r. spectroscopy. They contain either one or two neuraminic acid residues (alpha 2-->6)-linked to galactose in conventional biantennary N-acetyl-lactosamine-type glycans. Most contain fucose (alpha 1-->6)-linked to the terminal non-reducing N-acetylglucosamine.

scholarly journals Complementary DNA sequence of human amyloidogenic immunoglobulin light-chain precursors

1992 ◽  
Vol 285 (1) ◽  
pp. 149-152 ◽  
Author(s):  
P Aucouturier ◽  
A A Khamlichi ◽  
J L Preud'homme ◽  
M Bauwens ◽  
G Touchard ◽  
...  
Keyword(s):  
Amino Acid ◽  
Light Chain ◽  
Bone Marrow Cells ◽  
Northern Blotting ◽  
Glycosylation Site ◽  
Light Chains ◽  
Amino Acid Residues ◽  
Complementary Dna ◽  
Kidney Glomerulus ◽  
Complementarity Determining Regions

The primary structure of three amyloid precursor light chains was deduced from the sequence of complementary DNA (cDNA) from bone marrow cells from patients affected with classical lambda (patient Air) or kappa (patient Arn) amyloidosis and from a patient (Aub) in whom lambda amyloid deposits were unusual by their perimembranous location in the kidney glomerulus. All three RNAs were of normal size, as estimated by Northern blotting, and encoded normal-sized light chains. The deduced light-chain sequence from patient Arn was related to the V kappa 1 subgroup, and included ten residues that had not been previously reported at these positions, only one of which (Leu-21) was located in a beta-sheet (4-2). The unusual presence of Asn-70 determined a potential N-glycosylation site. The sequence of the light chain from patient Air belonged to the V lambda 1 subgroup, and included three unusually located amino acid residues, one of which had already been reported in an amyloidogenic lambda-chain. The sequence of the light chain from patient Aub was related to the V lambda 3 subgroup, and contained five amino acid residues that had not previously been described at the corresponding positions; two of them (His-36 and Ser-77) were located in beta-sheets (3-1 and 4-3 respectively). This sequence was also peculiar because of the presence of numerous acidic residues in the complementarity-determining regions. Such unusual primary structures might be responsible for the amyloidogenic properties of these light-chain precursors.

scholarly journals The amino acid sequence of rabbit J chain in secretory immunoglobulin A

1990 ◽  
Vol 271 (3) ◽  
pp. 641-647 ◽  
Author(s):  
G J Hughes ◽  
S Frutiger ◽  
N Paquet ◽  
J C Jaton
Keyword(s):  
Amino Acid ◽  
Immunoglobulin A ◽  
Complete Sequence ◽  
Glycosylation Site ◽  
Secretory Iga ◽  
Amino Acid Residues ◽  
J Chain ◽  
Mouse Sequence ◽  
Human Sequence ◽  
One Step

The primary structure of rabbit J chain, which occurs covalently bound to secretory IgA, was determined. J chain was isolated in its S-carboxymethylated form, in one step, by SDS/PAGE followed by electro-elution; 5 nmol of protein (approx. 75 micrograms), in all, was necessary for the determination of the complete sequence by the ‘shot-gun’ microsquencing technique; with the use of several site-specific endoproteinases, the various digests of S-carboxymethylated J chain were separated by micro-bore reverse-phase h.p.l.c. and the partial N-terminal sequences of all peptides were analysed. From the sequence alignment, gaps were filled by further extensive sequencing of the relevant overlapping fragments isolated from selected digests. Rabbit J chain comprises 136 amino acid residues, out of which eight are conserved cysteine residues, and is more closely similar to the human sequence (73.5% identify) than to the mouse sequence (68% identity). There is one unique glycosylation site at asparagine-48.

scholarly journals Mutational Insights into the Roles of Amino Acid Residues in Ligand Binding for Two Closely Related Family 16 Carbohydrate Binding Modules

2010 ◽  
Vol 285 (45) ◽  
pp. 34665-34676 ◽  
Author(s):  
Xiaoyun Su ◽  
Vinayak Agarwal ◽  
Dylan Dodd ◽  
Brian Bae ◽  
Roderick I. Mackie ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ligand Binding ◽  
Carbohydrate Binding ◽  
Amino Acid Residues ◽  
Related Family ◽  
Carbohydrate Binding Modules

scholarly journals Two pectate lyases from Caldicellulosiruptor bescii with the same CALG domain had distinct properties on plant biomass degradation

2020 ◽  
Author(s):  
Hamed I. Hamouda ◽  
Nasir Ali ◽  
Hang Su ◽  
Jie Feng ◽  
Ming Lu ◽  
...  
Keyword(s):  
Amino Acid ◽  
Plant Biomass ◽  
Degradation Products ◽  
Amino Acid Identity ◽  
Thermophilic Bacterium ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Acid Identity ◽  
Pectate Lyases ◽  
Caldicellulosiruptor Bescii

AbstractPectin deconstruction is the initial step in breaking the recalcitrance of plant biomass by using selected microorganisms that carry pectinolytic enzymes. Pectate lyases that cleave α-1,4-galacturonosidic linkage of pectin are widely used in industries, such as paper making and fruit softening. However, reports on pectate lyases with high thermostability are few. Two pectate lyases (CbPL3 and CbPL9) from a thermophilic bacterium Caldicellulosiruptor bescii were investigated. Although these two enzymes belonged to different families of polysaccharide lyase, both were Ca2+-dependent. Similar biochemical properties were shown under optimized conditions 80 °C–85 °C and pH 8–9. However, the degradation products on pectin and polygalacturonic acids (pGA) were different, revealing the distinct mode of action. A concanavalin A-like lectin/glucanase (CALG) domain, located in the N-terminus of two CbPLs, shares 100% amino acid identity. CALG-truncated mutant of CbPL9 showed lower activities than the wild-type, whereas the CbPL3 with CALG knock-out portion was reported with enhanced activities, thereby revealing the different roles of CALG in two CbPLs. I-TASSER predicted that the CALG in two CbPLs is structurally close to the family 66 carbohydrate binding module (CBM66). Furthermore, substrate-binding assay indicated that the catalytic domains in two CbPLs had strong affinities on pectate-related substrates, but CALG showed weak interaction with a number of lignocellulosic carbohydrates, except sodium carboxymethyl cellulose and sodium alginate. Finally, scanning electron microscope analysis and total reducing sugar assay showed that the two enzymes could improve the saccharification of switchgrass. The two CbPLs are impressive sources for degradation of plant biomass.ImportanceThermophilic proteins could be implemented in diverse industrial applications. We sought to characterize two pectate lyases, CbPL3 and CbPL9, from a thermophilic bacterium Caldicellulosiruptor bescii. The two enzymes had high optimum temperature, low optimum pH, and good thermostability at evaluated temperature. A family-66 carbohydrate binding module (CBM66) was identified in two CbPLs with sharing 100% amino acid identity. Deletion of CBM66 obviously decreased the activity of CbPL9, but increase the activity and thermostability of CbPL3, suggesting the different roles of CBM66 in two enzymes. Moreover, the degradation products by two CbPLs were different. These results revealed these enzymes could represent a potential pectate lyase for applications in paper and textile industries.

scholarly journals A Single Mutation in the Carbohydrate-Binding Module Enhances Cellulase Activity in Bacillus Amyloliquefaciens Mutant

2021 ◽  
Vol 18 (18) ◽  
Author(s):  
Nitipol POLSA ◽  
Chomphunuch SONGSIRIRITTHIGUL ◽  
Wasana SUYOTHA ◽  
Sugunya SUEBSAN ◽  
Somboon ANUNTALABHOCHAI ◽  
...  
Keyword(s):  
Amino Acid ◽  
Bacillus Amyloliquefaciens ◽  
Cellulase Activity ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Single Mutation ◽  
Plasma Technology ◽  
Lignocellulosic Wastes ◽  
Cellulose Binding ◽  
Hydrolysis Activity

From our earlier work, we modified the carbohydrate-binding module (CBM) of Bacillus amyloliquefaciens to increase cellulase activity using cold plasma technology. The cellulase gene (BglC-M) from the mutant was expressed in Escherichia coli BL21(DE3) under the T7 promoter. The hydrolysis activity of the cellulase mutant (BglC-M) was approximately 2.5-fold higher than the control (BglC-W) over a wide range of pH and temperature conditions. The amino acid sequence of the mutant BglC-M contained 471 residues that were almost identical to the control BglC-W. Only a single amino acid, lysine, was replaced by glutamic acid at position 370 (K370E) within the carbohydrate-binding module (CBM). Structure prediction and substrate docking of BglC-M indicated that the single mutation (K370E) might involve cellulose binding of the β-sandwich facilitated by hydrogen bonding. The docking study of cellopentaose with the model structure of BglC-M indicated that the replacement of lysine-370 led to the formation of a hydrogen bond with 436Y, which has a shorter distance (2.6 Å) compared with the control (5.4 Å). As a result, the structure becomes more compact and stable, resulting in increased catalytic efficiency. Finally, the biomass hydrolysis ability of cellulase was investigated on lignocellulosic wastes such as pineapple peel, corncob, and durian peel. The BglC-M enzyme showed a more significant amount of reducing sugar released from all lignocellulosic wastes than the control. This was the first evidence that altering the base composition of the cellulose binding module enhanced the catalytic activity. HIGHLIGHTS Increasing cellulase activity of Bacillus amyloliquefaciens using plasma technology Mutation at cellulose-binding module enhance cellulase hydrolysis activity Greater cellulase activity in the hydrolysis on lignocellulosic wastes GRAPHICAL ABSTRACT

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