Engineering of multiple arginines into the Ser/Thr surface of Trichoderma reesei endo-1,4-beta-xylanase II increases the thermotolerance and shifts the pH optimum towards alkaline pH

We studied the effects of increase in the number of surface arginines on the enzyme activity and stability of Trichoderma reesei endo-1,4-beta-xylanase II. The number of arginines was increased in two mutant series. The first set contained six arginines on different sides of the protein surface. These arginines had no significant effect on the thermostability. However, the optimal pH region became narrower. Another series of five arginines was engineered into the 'Ser/Thr surface', formed of part of the double-layered beta-sheet located on one side of the 'right-hand-like' xylanase. These mutations shifted the activity profile to the alkaline region by approximately 0.5-1.0 pH units. In addition, the arginines on the Ser/Thr surface increased the enzyme activity at high temperature, although the enzyme stability in the absence of substrate decreased significantly at 50-55 degrees C. In the presence of the substrate, the thermostability increased 4-5-fold at 60-65 degrees C. Thus, the substrate neutralized the destabilizing effect of Ser/Thr surface arginines and revealed a stabilizing effect of the same mutations. The stabilizing effect of arginines at high temperatures was seen clearly only when five arginines were introduced into the Ser/Thr surface.     

Protein-Metal Interactions Probed by SERS: Lysozyme on Nanostructured Gold Surface

Plasmonics - Surface-enhanced Raman scattering is a well-established technique for molecular detection at low concentration, which is becoming increasingly popular in the field of biotechnology and...     

Recording strategies and selection potential of feed intake measured using the X-ray method in rainbow trout

This study examines the way long-term feed intake should be recorded accurately for selective breeding purposes, and estimates selection potential in feed intake using the X-ray method to record individual daily feed intake in rainbow trout (Oncorhynchus mykiss). The analysis showed that the point estimates of daily feed intake displayed low repeatabilities (r = 0.09–0.32). This indicates that a minimum of three repeated records were needed to accurately record average feed intake at a fixed age. To effectively breed for feed intake over the whole growing period, it is necessary to determine average feed intake at different ages, since there were only moderate phenotypic and genetic correlations between average daily feed intake recorded at 140 g, 750 g and 2000 g wet mass. Heritability for average daily feed intake was low (average h² = 0.10), indicating that modest genetic changes can be obtained in response to selection. It was concluded that selection to genetically change long-term feed intake can be successful, yet repeated observations at several life stages are needed to ensure the accuracy of feed intake estimates and the efficiency of selection.     

Trade-off between warning signal efficacy and mating success in the wood tiger moth

The coloration of species can have multiple functions, such as predator avoidance and sexual signalling, that directly affect fitness. As selection should favour traits that positively affect fitness, the genes underlying the trait should reach fixation, thereby preventing the evolution of polymorphisms. This is particularly true for aposematic species that rely on coloration as a warning signal to advertise their unprofitability to predators. Nonetheless, there are numerous examples of aposematic species showing remarkable colour polymorphisms. We examined whether colour polymorphism in the wood tiger moth is maintained by trade-offs between different functions of coloration. In Finland, males of this species have two distinct colour morphs: white and yellow. The efficacy of the warning signal of these morphs was tested by offering them to blue tits in the laboratory. Birds hesitated significantly longer to attack yellow than white males. In a field experiment, the survival of the yellow males was also higher than white males. However, mating experiments in the laboratory revealed that yellow males had lower mating success than white males. Our results offer an explanation for the maintenance of polymorphism via trade-off between survival selection and mating success.     

Self-rated health as a predictor of disability retirement--the contribution of ill-health and working conditions

Objective

Self-rated health is a generic health indicator predicting mortality, many diseases, and need for care. We examined self-rated health as a predictor of subsequent disability retirement, and ill-health and working conditions as potential explanations for the association.

Methods

Self-rated health and the covariates were obtained from the Helsinki Health Study baseline mail surveys in 2000–2002 conducted among municipal employees aged 40–60 years (n = 6525). Data for disability retirement events (n = 625) along with diagnoses were linked from the Finnish Centre for Pensions, with a follow-up by the end of 2010. Hazard ratios (HR) and their 95% confidence intervals (CI) were calculated using competing risks models.

Results

Less than good self-rated health predicted disability retirement due to all causes among both women (HR = 4.60, 95% CI = 3.84–5.51) and men (HR = 3.83, 95% CI = 2.64–5.56), as well as due to musculoskeletal diseases (HR = 5.17, 95% CI = 4.02–6.66) and mental disorders (HR = 4.80, 95% CI = 3.50–6.59) among women and men pooled. Ill-health and physical working conditions partly explained the found associations, which nevertheless remained after the adjustments. Among the measures of ill-health limiting long-standing illness explained the association most in all-cause disability retirement and disability retirements due to musculoskeletal diseases, whereas common mental disorders explained the association most in disability retirements due to mental health disorders. Among working conditions physical work load and hazardous exposures at work explained the association most, although much less than ill-health.

Conclusions

Self-rated health is a strong predictor of disability retirement. This can be partly explained by ill-health and working conditions. Poor self-rated health provides a useful marker for increased risk of work disability and subsequent disability retirement.     

Elucidation of the Molecular Basis for Arabinoxylan-Debranching Activity of a Thermostable Family GH62 α-l-Arabinofuranosidase from Streptomyces thermoviolaceus

Xylan-debranching enzymes facilitate the complete hydrolysis of xylan and can be used to alter xylan chemistry. Here, the family GH62 α-l-arabinofuranosidase from Streptomyces thermoviolaceus (SthAbf62A) was shown to have a half-life of 60 min at 60°C and the ability to cleave α-1,3 l-arabinofuranose (l-Araf) from singly substituted xylopyranosyl (Xylp) backbone residues in wheat arabinoxylan; low levels of activity on arabinan as well as 4-nitrophenyl α-l-arabinofuranoside were also detected. After selective removal of α-1,3 l-Araf substituents from disubstituted Xylp residues present in wheat arabinoxylan, SthAbf62A could also cleave the remaining α-1,2 l-Araf substituents, confirming the ability of SthAbf62A to remove α-l-Araf residues that are (1→2) and (1→3) linked to monosubstituted β-d-Xylp sugars. Three-dimensional structures of SthAbf62A and its complex with xylotetraose and l-arabinose confirmed a five-bladed β-propeller fold and revealed a molecular Velcro in blade V between the β1 and β21 strands, a disulfide bond between Cys27 and Cys297, and a calcium ion coordinated in the central channel of the fold. The enzyme-arabinose complex structure further revealed a narrow and seemingly rigid l-arabinose binding pocket situated at the center of one side of the β propeller, which stabilized the arabinofuranosyl substituent through several hydrogen-bonding and hydrophobic interactions. The predicted catalytic amino acids were oriented toward this binding pocket, and the catalytic essentiality of Asp53 and Glu213 was confirmed by site-specific mutagenesis. Complex structures with xylotetraose revealed a shallow cleft for xylan backbone binding that is open at both ends and comprises multiple binding subsites above and flanking the l-arabinose binding pocket.     

The cultivation of oak seedlings inoculated with Tuber aestivum Vittad. in the boreal region of Finland

Despite recent findings, truffles are rarely found in Finland. In 2006, we began to explore the cultivation potential of Tuber aestivum/uncinatum in Finland. In 2006–2008, roughly 1,200 Quercus robur seedlings and 200 Q. pubescens seedlings were planted in 20 orchards. We aimed to challenge the Southern European (France) tree provenances of oak seedlings in a boreal climate. Additional winter coverings made up of fabric or plastic and twigs prevented the seedlings’ mortality even when the air temperature was below ?30 °C during the second winter. The results showed that the top soil temperature at 15 cm depth has to be above ?5 °C to guarantee the survival of seedlings. Q. pubescens was more sensitive to low soil temperatures than Q. robur. Morphological and PCR analysis of root samples collected over 2007–2010 confirmed the presence of T. aestivum in all orchards despite unfavorable temperatures during the winter time. The first T. aestivum sporocarps were found under Q. robur in October 2012 in the orchards established in 2006 on old agricultural land, showing truffle cultivation to be successful in the boreal climate.     

The effect of risedronate on osteogenic lineage is mediated by cyclooxygenase-2 gene upregulation

Introduction

The purpose of this study was to evaluate the effects of risedronate (Ris) in the modulation of bone formation in rats with glucocorticoid (GC)-induced osteoporosis by histomorphometric, immunohistochemical and gene expression analyses.

Methods

We analyzed structure, turnover and microarchitecture, cyclooxygenase 2 (COX-2) levels and osteocyte apoptosis in 40 female rats divided as follows: 1) vehicle of methylprednisolone (vGC) + vehicle of risedronate (vRis); 2) Ris 5 μg/Kg + vGC; 3) methylprednisolone (GC) 7 mg/Kg + vRis; 4) GC 7 mg/Kg +Ris 5 μg/Kg. In addition, we evaluated cell proliferation and expression of COX-2 and bone alkaline phosphatase (b-ALP) genes in bone marrow cells and MLO-y4 osteocytes treated with Ris alone or in co-treatment with the selective COX-2 inhibitor NS-398 or with dexametasone.

Results

Ris reduced apoptosis induced by GC of osteocytes (41% vs 86%, P < 0.0001) and increased COX-2 expression with respect to controls (Immuno-Hystochemical Score (IHS): 8.75 vs 1.00, P < 0.0001). These positive effects of Ris in bone formation were confirmed by in vitro data as the viability and expression of b-ALP gene in bone marrow cells resulted increased in a dose dependent manner.

Conclusions

These findings suggest a positive effect of Ris in bone formation and support the hypothesis that the up-regulation of COX-2 could be an additional mechanism of anabolic effect of Ris.     

Increased water resistance of CTMP fibers by oat (Avena sativa L.) husk lignin

The insertion of oat husk lignin onto chemithermomechanical pulp (CTMP) fibers was studied to increase fiber hydrophobicity. The pretreated pulp samples were subsequently used for preparation of handsheets for characterization. Treatment of CTMP with laccase in the presence of oat husk lignin resulted in a significant increase in hydrophobicity of the handsheet surface, as indicated by dynamic contact angle analysis. Water absorption time of 8 s was obtained with initial contact angle of 118°. Although the handsheet's brightness was reduced by 33%, tensile index was only subtly decreased. Neither laccase nor oat husk lignin alone gave much improved water absorption times. Therefore, handsheets made of laccase-treated pulp with and without oat husk lignin were further examined by XPS, which suggested that both laccase and oat husk lignin were inserted onto CTMP fibers. The oat husk lignin was distributed as heterogeneous aggregates on the handsheet surface whereas laccase was uniformly distributed. Evidence was obtained that the adsorbed laccase layer formed a noncovalent base for the insertion of oat husk lignin onto fiber surfaces.