Characterization of Polar and Non‐Polar Compounds of House Edible Bird's Nest (EBN) from Johor,Malaysia

This work investigated the polar (PC: protein, amino acid and metabolite) and non‐polar (NPC: fatty acid) compounds and bioactivity characteristics of the EBN harvested from the state of Johor in Malaysia. The electrophoretic gels exhibited 15 protein bands (16–173 kD) with unique protein profile. Amino acids analysis by AccQ?Tag method revealed 18 types of amino acids in EBN. Metabolite profiling was performed using High‐Performance Liquid Chromatography coupled with Quadrupole Time‐of‐Flight Mass Spectrometer (HPLC‐QTOF/MS) technique and a total of 54 compounds belonging to different groups were detected and identified. These findings help to uncover the relation of therapeutic activity of EBN. The EBN was further extracted with AcOEt and BuOH. The AcOEt extract was fractionated into three fractions (F₁?F₃), and the high triglyceride content in F₂ was verified by gC‐FID. The three groups of fatty acids discovered in EBN are 48.43 % of poly‐unsaturated (PUFA), 25.35 % of saturated fatty acids (SFA) and 24.74 % of mono‐unsaturated fat (MUFA). This is the first time to report results ofEBN, BuOH, and AcOEt extracts and of fraction F₂ (TEBN) on their analysis for their antioxidant activities by DPPH, ABTS and catalase assay and for their paraoxonase and anti‐tyrosinase activities. The results showed that TEBN exhibited the significant bioactivity in all assays. These findings suggest that TEBN is a good source for natural bioactive compounds in promoting body vigor. Current work widened the content of EBN especially on the triglyceride and also marked the content of specific location (Johor, Malaysia) of EBN origin.     

Expression and characterization of <Emphasis Type="Italic">Trichoderma virens</Emphasis> UKM-1 endochitinase in <Emphasis Type="Italic">Escherichia</Emphasis> <Emphasis Type="Italic">coli</Emphasis>

A gene encoding endochitinase from Trichoderma virens UKM-1 was cloned and expressed in E. coli BL21 (DE3). Both the endochitinase gene and its cDNA sequences were obtained. The endochitinase gene encodes 430 amino acids from an open reading frame comprising of 1,690 bp nucleotide sequence with three introns. The endochitinase was expressed as soluble and active enzyme at 20°C when induced with 1 mM IPTG. Maximum activity was observed at 4 h of post-induction time. SDS-PAGE showed that the purified endochitinase exhibited a single band with molecular weight of 42 kDa. Biochemical characterization of the enzyme displayed a near neutral pH characteristic with an optimum pH at 6.0 and optimum temperature at 50°C. The enzyme is stable between pH 3.0–7.0 and is able to retain its activity from 30 to 60°C. The presence of Mg²⁺ and Ca²⁺ ions increased the enzyme activity up to 20%. The purified enzyme has a strong affinity towards colloidal chitin and low effect on ethyl cellulose and D-cellubiose which are non-chitin related substrates. HPLC analysis from the chitin hydrolysis showed the release of (GlcNAc)₃, (GlcNAc)₂ and GlcNAc, in which (GlcNAc)₂ was the main product.     

Relief of Autoinhibition Enhances Vta1 Activation of Vps4 via the Vps4 Stimulatory Element

The endosomal sorting complexes required for transport (ESCRTs) impact multiple cellular processes including multivesicular body sorting, abscission, and viral budding. The AAA-ATPase Vps4 is required for ESCRT function, and its full activity is dependent upon the co-factor Vta1. The Vta1 carboxyl-terminal Vta1 SBP1 Lip5 (VSL) domain stimulates Vps4 function by facilitating oligomerization of Vps4 into its active state. Here we report the identification of the Vps4 stimulatory element (VSE) within Vta1 that is required for additional stimulation of Vps4 activity in vitro and in vivo. VSE activity is autoinhibited in a manner dependent upon the unstructured linker region joining the amino-terminal microtubule interacting and trafficking domains and the carboxyl-terminal VSL domain. The VSE is also required for Vta1-mediated Vps4 stimulation by ESCRT-III subunits Vps60 and Did2. These results suggest that ESCRT-III binding to the Vta1 microtubule interacting and trafficking domains relieves linker region autoinhibition of the VSE to produce maximal activation of Vps4 during ESCRT function.     

Bro1 stimulates Vps4 to promote intralumenal vesicle formation during multivesicular body biogenesis

Endosomal sorting complexes required for transport (ESCRT-0, -I, -II, -III) execute cargo sorting and intralumenal vesicle (ILV) formation during conversion of endosomes to multivesicular bodies (MVBs). The AAA-ATPase Vps4 regulates the ESCRT-III polymer to facilitate membrane remodeling and ILV scission during MVB biogenesis. Here, we show that the conserved V domain of ESCRT-associated protein Bro1 (the yeast homologue of mammalian proteins ALIX and HD-PTP) directly stimulates Vps4. This activity is required for MVB cargo sorting. Furthermore, the Bro1 V domain alone supports Vps4/ESCRT–driven ILV formation in vivo without efficient MVB cargo sorting. These results reveal a novel activity of the V domains of Bro1 homologues in licensing ESCRT-III–dependent ILV formation and suggest a role in coordinating cargo sorting with membrane remodeling during MVB sorting. Moreover, ubiquitin binding enhances V domain stimulation of Vps4 to promote ILV formation via the Bro1–Vps4–ESCRT-III axis, uncovering a novel role for ubiquitin during MVB biogenesis in addition to facilitating cargo recognition.     

The association between ownership of common household devices and obesity and diabetes in high,middle and low income countries

Background:

Household devices (e.g., television, car, computer) are common in high income countries, and their use has been linked to obesity and type 2 diabetes mellitus. We hypothesized that device ownership is associated with obesity and diabetes and that these effects are explained through reduced physical activity, increased sitting time and increased energy intake.

Methods:

We performed a cross-sectional analysis using data from the Prospective Urban Rural Epidemiology study involving 153 996 adults from high, upper-middle, lower-middle and low income countries. We used multilevel regression models to account for clustering at the community and country levels.

Results:

Ownership of a household device increased from low to high income countries (4% to 83% for all 3 devices) and was associated with decreased physical activity and increased sitting, dietary energy intake, body mass index and waist circumference. There was an increased odds of obesity and diabetes with the ownership of any 1 household device compared to no device ownership (obesity: odds ratio [OR] 1.43, 95% confidence interval [CI] 1.32–1.55; diabetes: OR 1.38, 95% CI 1.28–1.50). Ownership of a second device increased the odds further but ownership of a third device did not. Subsequent adjustment for lifestyle factors modestly attenuated these associations. Of the 3 devices, ownership of a television had the strongest association with obesity (OR 1.39, 95% CI 1.29–1.49) and diabetes (OR 1.33, 95% CI 1.23–1.44). When stratified by country income level, the odds of obesity and diabetes when owning all 3 devices was greatest in low income countries (obesity: OR 3.15, 95% CI 2.33–4.25; diabetes: OR 1.97, 95% CI 1.53–2.53) and decreased through country income levels such that we did not detect an association in high income countries.

Interpretation:

The ownership of household devices increased the likelihood of obesity and diabetes, and this was mediated in part by effects on physical activity, sitting time and dietary energy intake. With increasing ownership of household devices in developing countries, societal interventions are needed to mitigate their effects on poor health.The increasing global prevalence of obesity and type 2 diabetes mellitus has been driven predominantly by increases in high income countries.^1,2 However, increases are expected in low and middle income countries, due in part to rapid development and industrialization.Proximal determinants of obesity and diabetes include energy expenditure and intake;^3–5 however, the upstream factors are complex and entail numerous environmental factors. Of these, the increased use of common household devices (e.g., televisions, cars, computers) has been linked to increased sitting, decreased physical activity, obesity, metabolic syndrome and diabetes.^6–12 Time spent watching television has also been linked to poor diet¹³ and increased caloric intake.¹⁴ However, these findings are based on studies in high income countries where the ownership of these devices is common.^15,16 In low and middle income countries, such household devices are less prevalent, but their prevalence is rapidly increasing. Studies in countries with greater variability in the ownership of household devices are needed to understand the full effect of owning such devices on the risk of obesity and diabetes.We hypothesized that the ownership of a television, car or computer would be associated with an increased risk of obesity and diabetes and that these effects would be explained by reduced physical activity, increased sitting time and increased energy intake.     

Impact of incubation period on biodegradation of petroleum hydrocarbons from refinery wastewater in Kuantan,Malaysia by indigenous bacteria

This work studied the biodegradation of petroleum hydrocarbons (PHCs) extracted from refinery wastewater to produce industrially important by-products at different incubation periods. Two out of 13 bacterial isolates, KRD2 and KRA4 were isolated. Dichloromethane was used to extract the PHC, and gas chromatography-mass spectrometry (GC-MS) analysis revealed that the refinery wastewater PHC was successfully biodegraded using the selected bacterial isolates within 15 days of incubation. Both KRD2 and KRA4 isolates degraded all 13 initially extracted PHC compounds within 5 days, except C13BD and C9BD, which produced 6 and 4 compounds as secondary metabolites with peak area percentages of 1.58, 1.38, 0.85, 29.94, 7.59, and 11.16% and 3.55, 2.88, 52.31, and 6.14%, respectively. These metabolites have been reported in industrial and medical applications. After 10 days, only 6 and 8 compounds were degraded by both isolates, respectively, and C11PAD compound was produced, as well as C5PAD, C7PAD, and C13PAD. After 15 days, it was clear that all the initial PHC compounds have been completely degraded by both isolates. Metabolites C5PAD, C6PAD, C8PAD, and C13PAD were produced by KRD2, and metabolites C5PAD, C6PAD, C8PAD, and C9PAD were produced by KRA4 at different peak areas. The alignment revealed that the KRA4 isolate was included in the genus Chryseobacterium gambrini, while KRD2 isolate was successfully identified as Mycobacterium confluentis using the Biolog microbial identification system. The incubation period evidently affected biodegradation process by indigenous degraders. These effective bacteria were shown to be of great potential for further application in biodegradation technology of PHC contaminated refinery wastewater to produce industrially important by-products.     

RNAi-based transgene conferred extreme resistance to the geminivirus causing apical leaf curl disease in potato

Potato apical leaf curl disease is an emerging geminiviral disease in tropics and subtropics. It was reported for the first time in the year 1999 in northern plains of India but quickly spread to almost all potato growing regions of the country largely due to prevalence of warmer weather during early crop growth, thereby favoring whitefly vector. The problem of apical leaf curl disease in India became more severe due to lack of seed indexing for this virus in conventional seed production scheme. Although it accounts for major yield loss, there is no conventional source of resistance available in potato against Tomato Leaf Curl New Delhi Virus-Potato (ToLCNDV-Potato) that causes this disease in potato. In the present study, we have investigated the potential use of RNAi for obtaining resistance against this DNA virus in potato. The replication-associated protein gene (AC1) of the virus was used to obtain pathogen-derived resistance. The AC1 gene was PCR amplified from field-infected potato leaves, cloned and sequenced (JN393309). It showed 93% sequence similarity with the AC1 gene of Tomato Leaf Curl Virus-New Delhi (TOLCV-NDe; DQ169056) virus. Transgenic plants encoding the AC1 gene in three different orientations, viz. sense, antisense and hairpin loop, were raised. Transgenic lines when challenge inoculated with ToLCNDV-Potato showed different levels of resistance for all three constructs. Transgene integration and copy number in selected transgenic lines were determined by qPCR and further confirmed by Southern blot analysis. Though a reduction in viral titer was observed in transgenic lines encoding either antisense or hairpin loop constructs of AC1 gene, the latter transgenics showed most significant results as shown by reduction in the level of symptom expression in glasshouse screening as well as real-time data of in vivo virus concentration. In fact, we obtained a few totally asymptomatic transgenic lines with hairpin loop strategy.     

Exogenous proline significantly affects the plant growth and nitrogen assimilation enzymes activities in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis>) under salt stress

Salinity has been shown to be a major factor contributing to low nitrogen availability in plants. To verify the changes in nitrogen metabolism activity as affected by the exogenous application of proline under salt stress and its relation to salt tolerance, in vitro rice shoot apices were used as a model to study the growth performance and changes in nitrogen assimilation activities in two Malaysian rice cultivars MR 220 and MR 253. Results revealed that salt stress greatly reduced the plant height, shoot nitrate (NO₃ ^?) content, shoot glutamine synthetase (GS), and root nitrate reductase (NR) activities in both cultivars. Supplementation of proline significantly increased the plant height, number of roots, root NO₃ ^? content, root NR, and root GS activities under salt stress in both cultivars with greater enhancement in MR 253 than MR 220. The results also indicated that MR 253 possessed higher nitrite reductase (NiR) and glutamate synthase (NADH–GOGAT) activities as compared with MR 220 in all tested treatments. It was suggested that the NO₃ ^? content, NR, and GS activities played important roles in regulating nitrogen metabolism under salt stress. Taken together, it was concluded that the ability of proline in mitigating salt stress-induced damages was correlated with the changes in nitrogen assimilation activities.