A proteomic approach to study Cry1Ac binding proteins and their alterations in resistant Heliothis virescens larvae

Binding of the Bacillus thuringiensis Cry1Ac toxin to specific receptors in the midgut brush border membrane is required for toxicity. Alteration of these receptors is the most reported mechanism of resistance. We used a proteomic approach to identify Cry1Ac binding proteins from intestinal brush border membrane (BBM) prepared from Heliothis virescens larvae. Cry1Ac binding BBM proteins were detected in 2D blots and identified using peptide mass fingerprinting (PMF) or de novo sequencing. Among other proteins, the membrane bound alkaline phosphatase (HvALP), and a novel phosphatase, were identified as Cry1Ac binding proteins. Reduction of HvALP expression levels correlated directly with resistance to Cry1Ac in the YHD2-B strain of H. virescens. To study additional proteomic alterations in resistant H. virescens larvae, we used two-dimensional differential in-gel electrophoresis (2D-DIGE) to compare three independent resistant strains with a susceptible strain. Our results validate the use of proteomic approaches to identify toxin binding proteins and proteome alterations in resistant insects.     

Detection of protein adduction derived from styrene oxide to cysteine residues by alkaline permethylation

Styrene oxide-cysteine adduction is predominantly involved in protein covalent modification after exposure in vivo to styrene or styrene oxide. In the present study, we developed an alkaline permethylation- and GC/MS-based approach to detect styrene oxide-derived protein adduction. Permethylation of the protein adducts produced two methylthiophenylethanols, namely 2-methylthio-2-phenyl-1-ethanol and 2-methylthio-1-phenyl-1-ethanol. To improve the permethylation efficiency, reaction conditions, including temperature, time, NaOH strength, and molar ratio of CH₃I/NaOH, were explored. Under optimized conditions, the yields of the analyte formation resulting from permethylation of authentic standard α- and β-mercapturic acids, representing α and β isomers of cysteine adducts, were 35% and 28%, respectively. Permethylation of styrene oxide-modified bovine serum albumin released the two methylthiophenylethanols with an α-/β-adduction ratio of 1.5. A concentration-dependent increase in both α- and β-adduction was observed in mouse liver microsomes incubated with styrene at various concentrations. CD-1 mice were administered intraperitoneally with styrene at doses of 0, 50, and 400 mg/kg daily for 5 days. The formation of protein adducts derived from styrene oxide in whole blood in 400 mg/kg group was observed with an α/β ratio of 4.8, suggesting that the reaction of styrene oxide with cysteine residues took place more likely at the α-carbon than the β-carbon of styrene oxide.     

An organic solvent-stable alkaline protease from Pseudomonas aeruginosa strain K: Enzyme purification and characterization

The organic solvent-tolerant strain K protease was purified to homogeneity by ammonium sulphate precipitation and anion exchange chromatography with 124-fold increase in specific activity. The molecular mass of the purified enzyme as revealed by SDS-PAGE electrophoresis is 51,000 Da. The strain K protease was an alkaline metalloprotease with an optimum pH and temperature of 10 and 70 °C, respectively. The enzyme showed stability and activation in the presence of organic solvents with log P_a/w values equal or more than 4.0. After 14 days of incubation, the purified protease was activated 1.11, 1.82, 1.50, 1.75 and 1.80 times in 1-decanol, isooctane, decane, dodecane and hexadecane, respectively.     

Association between alkaline phosphatase and hypertension in a rural Japanese population: The Nagasaki Islands study

Background

Although serum alkaline phosphatase (ALP) levels have been associated with hypertension, and ALP is known as an enzyme affected by alcohol consumption, no study has been published on the associations between ALP and the risk of hypertension in relation to drinking status.

Methods

We conducted a cross-sectional study of 2,681 participants (837 men and 1,846 women) aged 30 to 89 years undergoing a general health check-up to investigate the associations between ALP and hypertension in relation to drinking status.

Results

Of the 2,681 participants, 1,549 (514 men and 1,035 women) were diagnosed with hypertension. A sex difference was observed for the relationship between ALP and hypertension. While no significant association was observed for men, the association was significantly positive for women. The multivariable adjusted odds ratio and 95% coincidence interval (CI) of hypertension per increment of 1-log ALP were 0.95 (95% CI: 0.56 to 1.59) for men and 1.57 (95% CI: 1.07 to 2.33) for women. When this analysis was restricted to nondrinkers, a significantly elevated risk of hypertension was observed for men and remained significant for women; that is, 3.32 (95% CI: 1.38 to 8.02) for men and 1.68 (95% CI: 1.11 to 2.55) for women.

Conclusion

ALP is associated with hypertension for both male and female nondrinkers, but not for drinkers. For analyses of associations between ALP and blood pressure, alcohol consumption should thus be considered a potential confounder.     

Alkaline phosphatase-to-albumin ratio as a novel predictor of long-term adverse outcomes in coronary artery disease patients who underwent PCI

Background: Alkaline phosphatase (ALP) and albumin (ALB) have been shown to be associated with coronary artery disease (CAD), and it has been reported that alkaline phosphatase-to-albumin ratio (AAR) is associated with the liver damage and poorer prognosis of patients with digestive system malignancy. Moreover, several previous studies showed that there was a higher incidence of malignancy in CAD patients. However, to our knowledge, the relationship between AAR and long-term adverse outcomes in CAD patients after undergoing percutaneous coronary intervention (PCI) has not been investigated. Therefore, we aim to access the relation between AAR and long-term adverse outcomes in post-PCI patients with CAD.Methods: A total of 3378 post-PCI patients with CAD were enrolled in the retrospective Clinical Outcomes and Risk Factors of Patients with Coronary Heart Disease after PCI (CORFCHD-ZZ) study from January 2013 to December 2017. The median duration of follow-up was 37.59 ± 22.24 months. The primary end point was long-term mortality including all-cause mortality (ACM) and cardiac mortality (CM). The secondary end points were major adverse cardiac events (MACEs) and major adverse cardiac and cerebrovascular events (MACCEs).Results: Kaplan–Meier analyses showed that an increased AAR was positively correlated with incidences of long-term ACM (log-rank, P=0.014), CM (log-rank, P=0.011), MACEs (log-rank, P=0.013) and MACCEs (log-rank, P=0.006). Multivariate Cox regression analyses showed that the elevated AAR was an independent predictor of long-term ACM (adjusted HR = 1.488 [1.031–2.149], P=0.034), CM (adjusted HR = 1.837 [1.141–2.959], P=0.012), MACEs (adjusted HR = 1.257 [1.018–1.551], P=0.033) and MACCEs (adjusted HR = 1.237 [1.029–1.486], P=0.024).Conclusion: An elevated AAR is a novel independent predictor of long-term adverse outcomes in CAD patients following PCI.     

The unmasking of lignin structures in wheat straw by alkali

This study reports on the structural modifications of wheat straw cell wall promoted by potassium carbonate and sodium hydroxide that lead to the unmasking of some lignin structures. The first impact of the treatments was the extraction of a particular fraction of lignin enriched in C-C linked structures compared to the mean composition in reference wheat straw. Concomitantly, an apparent increase in the amount of lignin monomers released by the cleavage of alkyl-aryl ether bonds was observed in alkali-extracted samples. By summing the amount of ether linked monomers analyzed by thioacidolysis in the solubilized lignin to that found in the extracted wheat straw, an excess of up to 37% is apparent, relative to the corresponding amount in the reference wheat straw. Other modifications of the cell wall were also found. Indeed, a fraction of uronic acids was lost during the treatments and a new fractionation pattern of the lignin-carbohydrate complexes was evidenced. It can thus be concluded that a significant proportion of lignin within the cell wall was unmasked after (i) the selective removal of a particular lignin fraction, (ii) a partial saponification of the esterified fraction of lignin with uronic acids and (iii) a modification of the interactions between the cell wall constituents.     

水稻发芽期和幼苗前期耐碱性的鉴定方法研究

通过采用4种不同浓度的Na2CO3模拟碱胁迫,以水稻发芽期发芽势和发芽率的相对碱害率以及幼苗前期的根数、根长和苗高的相对碱害率为鉴定评价指标,研究水稻发芽期和幼苗前期耐碱性鉴定的碱胁迫条件。结果表明,水稻发芽势对碱胁迫的反应较发芽率更为敏感;根长对碱胁迫的反应较苗高和根数更为敏感;幼苗前期对碱胁迫的反应较发芽期更为敏感。在0．15％或0．20％Na2CO3胁迫下水稻发芽势和发芽率的相对碱害率平均值、方差和变异幅度较大。且其分布均匀合理,可作为水稻发芽期耐碱性鉴定的碱胁迫条件;而在0．10％或0．15％Na2CO3胁迫下,幼苗前期根数、根长和苗高的相对碱害率以及综合相对碱害率的平均值、方差和变异幅度较大,且分布相对均匀合理,可作为水稻幼苗前期耐碱性鉴定的碱胁迫条件。在碱胁迫下,水稻发芽势、发芽率、根数、根长和苗高的表型值分别与它们自身的相对碱害率呈极显著的负相关;发芽率的相对碱害率与根数、根长和苗高的相对碱害率呈显著的正相关;发芽期的综合相对碱害率与幼苗前期的综合相对碱害率呈显著的正相关;因此,以水稻发芽期的耐碱性可间接判断水稻幼苗前期的耐碱性。     

Germination variability and the effect of various pre-treatment on germination in the perennial spurge Euphorbia nicaeensis All.

Patterns of seed germination of the perennial spurge Euphorbia nicaeensis were studied in three populations in south-western Spain. We investigated the variation in seed viability and germination among individuals, and among populations over 2 consecutive years. We also studied if diverse factors such as temperature, acid scarification, darkness or caruncle loss affect the germination of the seeds.

Interindividual and interpopulation variability in seed viability was found. E. nicaeensis seeds are nondormant, so differences in interpopulation viability translate into differences in final germination rates. The germination percentage of the seeds from each population was similar in the 2 years studied. The effects of diverse factors were homogeneous in the two populations studied. Darkness has no effect on seed germination, and ecarunculate seeds germinated in the same proportions as carunculate seeds; this could allow the seeds to germinate in the chambers of deserted anthills. Acid scarification significantly reduced the germination percentage in only one of the populations, but over 50% of the seeds germinated, which could allow herbivores to act as occasional dispersing agents. The seeds that were preheated at 100 °C for 1 and 5 min germinated in the same proportions as the control group. The seeds that were preheated at 120 °C for 5 min displayed a significant decrease in germination, but the percentage was over 40% for both populations, indicating that the seeds could still germinate after the passage of a fire.     

Sambhar Salt Lake

The saline and alkaline brines from the Sambhar Salt Lake (SSL), both from the main lake and from the solar evaporation pans at Sambhar Salt Limited, Sambhar, Rajasthan, India, were studied with respect to their chemical composition and presence of red, extremely haloalkaliphilic archaebacteria. The brines had pH values of 9.5±0.2 and a total salt content ranging from 7% (w/v) to more than 30% (w/v). Sodium chloride, sodium carbonate, sodium bicarbonate and sodium sulphate were the principal salts present in these brines which lacked divalent cations (calcium and magnesium). Six strains of red, extremely haloalkaliphilic bacteria, designated SSL 1 to SSL 6, were isolated. All the isolates showed obligate requirements for sodium chloride (>15%, w/v) and high pH (>9.0). Magnesium ions were required in traces for maintaining morphological structure and pigmentation. All these strains possessed the diether core lipids, phosphatidylglycerol (PG), phosphatidylglycerophosphate (PGP), and bacterioruberins characteristic of halophilic archaebacteria. The strains were assigned to the newly proposed genus Natronobacterium.Part of the paper was presented by the authors at XIV International Congress of Microbiology 7–13 September 1986, Manchester, UK