Recent developments on cellulases and carbohydrate-binding modules with cellulose affinity

This review concerns basic research on cellulases and cellulose-specific carbohydrate-binding modules (CBMs). As a background, glycosyl hydrolases are also briefly reviewed. The nomenclature of cellulases and CBMs is discussed. The main cellulase-producing organisms and their cellulases are described. Synergy, enantioseparation, cellulases in plants, cellulosomes, cellulases and CBMs as analytical tools and cellulase-like enzymes are also briefly reviewed.     

Generation of ligand-receptor alliances by "SEA" module-mediated cleavage of membrane-associated mucin proteins

A mechanism is described whereby one and the same gene can encode both a receptor protein as well as its specific ligand. Generation of this receptor-ligand partnership is effected by proteolytic cleavage within a specific module located in a membrane resident protein. It is postulated here that the "SEA" module, found in a number of heavily O-linked glycosylated membrane-associated proteins, serves as a site for proteolytic cleavage. The subunits generated by proteolytic cleavage of the SEA module reassociate, and can subsequently elicit a signaling cascade. We hypothesize that all membrane resident proteins containing such a "SEA" module will undergo cleavage, thereby generating a receptor-ligand alliance. This requires that the protein subunits resulting from the proteolytic cleavage reassociate with each other in a highly specific fashion. The same SEA module that serves as the site for proteolytic cleavage, probably also contains the binding sites for reassociation of the resultant two subunits. More than one type of module can function as a site for proteolytic cleavage; this can occur not only in one-pass membrane proteins but also in 7-transmembrane proteins and other membrane-associated proteins. The proposal presented here is likely to have significant practical consequences. It could well lead to the rational design and identification of molecules that, by binding to one of the cleaved partners, will act either as agonists or antagonists, alter signal transduction and, hence, cellular behavior.     

De novo prediction of three-dimensional structures for major protein families

The family 10 xylanase from Streptomyces olivaceoviridis E-86 contains a (beta/alpha)(8)-barrel as a catalytic domain, a family 13 carbohydrate binding module (CBM) as a xylan binding domain (XBD) and a Gly/Pro-rich linker between them. The crystal structure of this enzyme showed that XBD has three similar subdomains, as indicated by the presence of a triple-repeated sequence, forming a galactose binding lectin fold similar to that found in the ricin toxin B-chain. Comparison with the structure of ricin/lactose complex suggests three potential sugar binding sites in XBD. In order to understand how XBD binds to the xylan chain, we analyzed the sugar-complex structure by the soaking experiment method using the xylooligosaccharides and other sugars. In the catalytic cleft, bound sugars were observed in the xylobiose and xylotriose complex structures. In the XBD, bound sugars were identified in subdomains alpha and gamma in all of the complexes with xylose, xylobiose, xylotriose, glucose, galactose and lactose. XBD binds xylose or xylooligosaccharides at the same sugar binding sites as in the case of the ricin/lactose complex but its binding manner for xylose and xylooligosaccharides is different from the galactose binding mode in ricin, even though XBD binds galactose in the same manner as in the ricin/galactose complex. These different binding modes are utilized efficiently and differently to bind the long substrate to xylanase and ricin-type lectin. XBD can bind any xylose in the xylan backbone, whereas ricin-type lectin recognizes the terminal galactose to sandwich the large sugar chain, even though the two domains have the same family 13 CBM structure. Family 13 CBM has rather loose and broad sugar specificities and is used by some kinds of proteins to bind their target sugars. In such enzyme, XBD binds xylan, and the catalytic domain may assume a flexible position with respect to the XBD/xylan complex, inasmuch as the linker region is unstructured.     

Can we infer peptide recognition specificity mediated by SH3 domains?

Protein interaction domain families that modulate the formation of macromolecular complexes recognize specific sequence or structural motifs. For instance SH3 and WW domains bind to polyproline peptides while SH2 and FHA domains bind to peptides phosphorylated in Tyr and Thr respectively. Within each family, variations in the chemical characteristics of the domain binding pocket modulate a finer peptide recognition specificity and, as a consequence, determine the selection of functional protein partners in vivo. In the proteomic era there is the need for reliable inference methods to help restricting the sequence space of the putative targets to be confirmed experimentally by more laborious experimental approaches. Here we will review the published data about the peptide recognition specificity of the SH3 domain family and we will propose a classification of SH3 domains into eight classes. Finally, we will discuss whether the available information is sufficient to infer the recognition specificity of any uncharacterized SH3 domain.     

Biochemical characterization of Thermotoga maritima endoglucanase Cel74 with and without a carbohydrate binding module (CBM)

The genome of the hyperthermophilic bacterium Thermotoga maritima (Tm) encodes at least eight glycoside hydrolases with putative signal peptides; the biochemical characteristics of seven of these have been reported previously. The eighth, Tm Cel74, is encoded by an open reading frame of 2124 bp corresponding to a polypeptide of 79 kDa with a signal peptide at the amino-terminus. The gene (lacking the signal peptide) encoding Tm Cel74 was expressed as a 77 kDa monomeric polypeptide in Escherichia coli and found to be optimally active at pH 6, 90 degrees C, with a melting temperature of approximately 105 degrees C. The cel74 gene was previously found to be induced during T. maritima growth on a variety of polysaccharides, including barley glucan, carboxymethyl cellulose (CMC), glucomannan, galactomannan and starch. However, while Tm Cel74 was most active towards barley glucan and to a lesser extent CMC, glucomannan and tamarind (xyloglucan), no activity was detected on other glycans, including galactomannan, laminarin and starch. Also, Tm Cel74 did not contain a carbohydrate binding module (CBM), versions of which have been identified in the amino acid sequences of other family 74 enzymes. As such, a CBM associated with a chitinase in another hyperthermophile, Pyrococcus furiosus, was used to create a fusion protein that was active on crystalline cellulose; Tm Cel74 lacked activity on this substrate. Based on the cleavage pattern determined for Tm Cel74 on glucan-based substrates, this enzyme likely initiates recruitment of carbohydrate carbon and energy sources by creating oligosaccharides that are transported into the cell for further processing.     

羊草无性系构件在不同环境下的可塑性变化

对温室内栽培羊草,人为控制温度及光照强度的条件,观测无性系构件的生长与分化的结果表明,羊草的无性系构件生长存在明显的可塑性变化,长时间的低温显著抑制羊草根茎和无性小株的分化（Ｐ＜０．０５）,而对分蘖无明显抑制作用．遮荫显著抑制根茎和无性小株的分化（Ｐ＜０．０５）,但对分蘖的影响并不显著．羊草的这种生态适应特征有利于对环境变化的适应．     

嗜冷嗜酸β-1,4-木聚糖酶及其钙离子依赖型碳水化合物结合模块

【目的】β-1,4-木聚糖酶是木聚糖降解的关键酶之一,嗜冷嗜酸木聚糖酶在功能性低聚木糖的制备中具有重要作用,但相关报道较少。【方法】从太平洋火色杆菌(Flammeovirga pacifica)菌株WPAGA1基因组发掘到一条新型的木聚糖酶序列,经基因合成、质粒构建和表达,并对其进行分离纯化及酶学性质研究。【结果】该木聚糖酶(Xyl4513)具有2个保守结构域,一个属于糖苷水解酶11家族(glycoside hydrolase family 11,GH11)催化模块(Xyl4513-T),另一个属于碳水化合物结合模块(carbohydrate-binding module,CBM) 60家族(CBM4513),这是一种非常罕见的GH11家族木聚糖酶含有CBM的现象。纯化后的Xyl4513最适反应温度和pH值分别为30℃、3.0,这一特性说明Xyl4513为嗜冷嗜酸β-1,4-木聚糖酶;而截短的木聚糖酶Xyl4513-T最适反应温度和pH值分别为20℃、4.0,且催化效率(k_cat/K_m)较前者下降了20%,说明CBM4513对酶稳定性和催化效...     

The occurrence and effectiveness of hypersensitive reaction against galling herbivores across host taxa

1. Hypersensitive reaction is an important type of induced defence by which the plant elicits a defence response to pathogens and insects. Hypersensitive reaction has been argued to be the most common plant resistance mechanism against insect herbivores that have intimate associations with their host plants. 2. The work reported here attempted to establish how important and widespread hypersensitive reaction might be against gall‐forming species across host taxa. 3. Hypersensitive reaction was the most important mortality factor against gall formation across host plant taxa in seven out of eight cases. 4. The number of insect galls correlated with the size of the leaves but module (leaf) size was a weak factor influencing the incidence of plant hypersensitive reaction to galling. 5. Insect galls and hypersensitive reactions occurred in genetically distant as well as geographically widespread host plant taxa.