Crystal structure of a papain-E-64 complex

E-64 [1-[N-[(L-3-trans-carboxyoxirane-2-carbonyl)-L-leucyl] amino]-4-guanidinobutane] is an irreversible inhibitor of many cysteine proteases. A papain-E-64 complex was crystallized at pH 6.3 by using the hanging drop method. Three different crystal forms grew in 3-7 days; the form chosen for structure analysis has space group P212121, with a = 42.91(4) A, b = 102.02(6) A, c = 49.73(2) A, and Z = 4. Diffraction data were measured to 2.4-A resolution, giving 9367 unique reflections. The papain structure was solved by use of the molecular replacement method, and then the inhibitor was located from a difference electron density map and fitted with the aid of a PS330 computer graphics system. The structure of the complex was refined to R = 23.3%. Our analysis shows that a covalent link is formed between the sulfur of the active-site cysteine 25 and the C-2 atom of the inhibitor. Contrary to earlier predictions, the E-64 inhibitor clearly interacts with the S subsites on the enzyme rather than the S' subsites, and papain's histidine 159 imidazole group plays a binding rather than a catalytic role in the inactivation process.     

Role of nitrogen and magnesium for growth,yield and nutritional quality of radish

Nitrogen (N) affects all levels of plant function from metabolism to resource allocation, growth, and development and Magnesium (Mg) is a macronutrient that is necessary to both plant growth and health. Radish (Raphanus sativus L.) occupies an important position in the production and consumption of vegetables globally, but there are still many problems and challenges in its nutrient management. A pot trial was conducted to investigate the effects of nitrogen and magnesium fertilizers on radish during the year 2018–2019. Nitrogen and magnesium was applied at three rates (0, 0.200, and 0.300 g N kg⁻¹ soil) and (0, 0.050, and 0.100 g Mg kg⁻¹ soil) respectively. The experiment was laid out in a completely randomized design (CRD) and each treatment was replicated three times. Growth, yield and quality indicators of radish (plant height, root length, shoot length, plant weight, total soluble sugar, ascorbic acid, total soluble protein, crude fiber, etc.) were studied. The results indicated that different rates of nitrogen and magnesium fertilizer not only influence the growth dynamics and yields but also enhances radish quality. The results revealed that the growth, yield and nutrient contents of radish were increased at a range of 0.00 g N. kg⁻¹ soil to 0.300 g N. kg⁻¹ soil and 0.00 g Mg. kg⁻¹ soil to 0.050 g Mg. kg⁻¹ soil and then decreased gradually at a level of 0.100 g Mg. kg⁻¹ soil. In contrast, the crude fiber contents in radish decreased significantly with increasing nitrogen and magnesium level but increased significantly at Mg₂ level (0.050 g Mg. kg⁻¹ soil). The current study produced helpful results for increasing radish quality, decreasing production costs, and diminishing underground water contamination.     

Modern and ancestral genotypes of Mycobacterium tuberculosis from Andhra Pradesh, India

Traditionally, the distribution of the Mycobacterium tuberculosis genotypes in India has been characterized by widespread prevalence of ancestral lineages (TbD1+ strains and variants) in the south and the modern forms (TbD1(-) CAS and variants) predominating in the north of India. The pattern was, however, not clearly known in the south-central region such as Hyderabad and the rest of the state of Andhra Pradesh where the prevalence of both tuberculosis (TB) and human immunodeficiency virus (HIV) infection is one of the highest in the country; this area has been the hotspot of TB vaccine trials. Spoligotyping of 101 clinical isolates obtained from Hyderabad and rural Andhra Pradesh confirmed the occurrence of major genogroups such as the ancestral (or the TbD1+ type or the East African Indian (EAI) type), the Central Asian (CAS) or Delhi type and the Beijing lineage in Andhra Pradesh. Sixty five different spoligotype patterns were observed for the isolates included in this study; these were further analyzed based on specific genetic signatures/mutations. It was found that the major genogroups, CAS and "ancestral," were almost equally prevalent in our collection but followed a north-south compartmentalization as was also reported previously. However, we observed a significant presence of MANU lineage in south Andhra Pradesh, which was earlier reported to be overwhelmingly present in Mumbai. This study portrays genotypic diversity of M. tuberculosis from the Indian state of Andhra Pradesh and provides a much needed snapshot of the strain diversity that will be helpful in devising effective TB control programs in this part of the world.     

Allergenic Implications of Airborne Leptosphaeria Ascospores

In order to investigate potential acroallergens of the Auckland region, a quantitative and qualitative study of the air spora was conducted at three different sites over the 12-month period, September 1979 to August 1980. Burkard volumetric spore traps were operated simultaneously at each of the three sites viz. Mt. Eden, Henderson and Waitakeres, located on a 20-km axis extending westward from Auckland City and encompassing residential, horticultural, agricultural, commercial and forested contexts. In the individual category of ascomycetous fungi, Leptosphaeria ascospores were recorded as an important component of air spora of the two non-forested sites. Ascospores concentration displayed a seasonal peak in late summer (February) and early autumn (March) and a diel periodicity with a distinct nocturnal maxima at all sites, confirming Leptosphaeria to be a component of the “rising air” or “damp-air spora”. The maximum concentration exceeded 4430 m^?3 of air around midnight (24/25 March, early autumn). A comparison of the results from the three sites showed that 61% of the total Leptosphaeria ascospores were trapped at Henderson (a satellite town in a rural setting) 29% at Mt. Eden (commercial/residential area) and 10% at Waitakeres (mainly forest site). Our data for Leptosphaeria ascospores combined with a high regional incidence of respiratory allergic diseases particularly bronchial asthma in late summer and autumn, indicate that a thorough investigation of the role of Leptosphaeria as a potential aeroallergen is warranted.     

Interactions of bacterial cytokinins and IAA in the rhizosphere may alter phytostimulatory efficiency of rhizobacteria

Phytohormones from rhizobacterial origin have been linked to their phytostimulation potential. However, while studying the efficacy of plant growth promoting bacteria, focus has always been on a single hormone. The role of plant hormones often overlay and they mutually modulate their effect. In current study focus was on the role of two hormones (cytokinins and indole acetic acid) in phytostimulation by rhizobacteria. Endogenous rhizosphere bacteria were isolated and screened for the presence of phytohormones. Bacterial strains from three different genera (Pseudomonas, Bacillus and Azospirillum) were screened positive for cytokinins and IAA. Phytohormones were simultaneously determined in SPE purified bacterial extract by ultra performance liquid chromatography (UPLC) coupled to a tandem mass spectrometer through electrospray interface. Cytokinins and IAA were determined in positive and negative mode, respectively with MRM scan. Zeatin, zeatin riboside and dihydrozeatin riboside were detected and quantified in the selected strains. Significant positive correlation between cytokinins and IAA in bacterial culture and plant endogenous hormones (r = 0.933 and r = 0.983; P = 0.01, respectively) was observed. However, strains with high IAA to cytokinins ratio could hardly enhance in-planta cytokinins, indicating antagonistic relation between the two hormones. Significant correlation of cytokinin with shoot length (r = 0.797; P = 0.01), fresh weight (r = 0.685; P = 0.01) and dry weight (r = 0.704; P = 0.01) was reported under axenic conditions. Bacterial IAA was correlated negatively to root length (r = 0.853; P = 0.01) and positively correlated to the number of roots (r = 0.964; P = 0.01). In natural conditions maximum increase in spike length (33%), number of tillers (71%) and weight of seeds (39%) was documented at final harvest in bacterially inoculated plants.     

Potential of bacterial indoleacetic acid to induce adventitious shoots in plant tissue culture

AIMS: The main aim of this study was to investigate the possible role of indoleacetic acid (IAA) from bacteria to induce in vitro adventitious shoots in internodal explants of Brassica oleracea L. METHODS AND RESULTS: Culture supernatant of Halomonas sp. RE1 and Halomonas sp. HT1 that contain 21 and 40 microg ml(-1) IAA, respectively, was used to supplement Murashige and Skoog (MS) medium. Two combinations that were supplemented with bacterial supernatant (BS) are MS + BS and MS + BS + 10%CW (coconut water) while basal MS medium was used as control. The amounts of BS used in this experiment were 50, 100, 150 and 200 microl in 5 ml MS medium in each combination. In vitro-grown internodal explants of B. oleracea were inoculated on these media combinations and incubated in a growth chamber at 25 +/- 1 degrees C and exposed to 16-h cool fluorescent light. After 5-6 weeks of incubation adventitious shoot induction was observed in all treatments that were supplemented with BS as compared with the controls where very low response was observed. The frequency of shoot induction was high in media that were supplemented with 10%CW in the presence of bacterial auxin. CONCLUSIONS: It was concluded that IAA of microbial origin has the potential to induce adventitious shoots in internodal explants. SIGNIFICANCE AND IMPACT OF THE STUDY: IAA from bacteria can be effectively used in plant tissue culture; especially a combination of MS + BS + 10%CW is very cost-effective as compared with synthetic phytohormones for in vitro studies.