Antifungal activity of mangrove rhizobacterium Pseudomonas aeruginosa against certain phytopathogenic fungi and its growth characterization

Antimicrobial substances are widespread and they are likely to play an important protective role. Marine bacterium has been recognized as producer of important antimicrobial substances which has an exceedingly bright future in the discovery of life saving drugs. The present study was carried out to screen the antifungal activity of mangrove rhizobacteria against certain phyto pathogens from Manakudi estuary, Kanyakumari District, Tamilnadu. Around 20 colonies obtained in Zobell marine agar plates were screened for antifungal traits. Among the 20 isolates, the candidate bacterial isolate exhibited good anti fungal ability. Identification of strains was carried out and confirmed by cultural, biochemical and 16S rDNA sequences. The potent strain was identified as Pseudomonas aeruginosa. Various process factors such as different pH, temperature, carbon and nitrogen sources and NaCl were tested for the bacterial growth in static and shaking conditions. The isolated Pseudomonas aeruginosa possesed a variety of promising properties that favoured as a better biocontrol agent. In the present investigation antifungal activity of the mangrove isolate was tested against common pathogens like Penicillium sp., Candida sp., Aspergillus sp., Aspergillus fumigatus, Aspergillus flavus, Pescalotionbsis sp., Fusarium oxysporum and Glomerella cinculata. The candidate bacterium showed inhibitory action to the tested fungal pathogens except Fusarium oxysporum and Glomerella cinculata.

HspB2/Myotonic Dystrophy Protein Kinase Binding Protein (MKBP) as a Novel Molecular Chaperone: Structural and Functional Aspects

The small heat shock protein, human HspB2, also known as Myotonic Dystrophy Kinase Binding Protein (MKBP), specifically associates with and activates Myotonic Dystrophy Protein Kinase (DMPK), a serine/threonine protein kinase that plays an important role in maintaining muscle structure and function. The structure and function of HspB2 are not well understood. We have cloned and expressed the protein in E.coli and purified it to homogeneity. Far-UV circular dichroic spectrum of the recombinant HspB2 shows a b-sheet structure. Fluorescence spectroscopic studies show that the sole tryptophan residue at the 130 th position is almost completely solvent-exposed. Bis-ANS binding shows that though HspB2 exhibits accessible hydrophobic surfaces, it is significantly less than that exhibited by another well characterized small HSP, aB-crystallin. Sedimentation velocity measurements show that the protein exhibits concentration-dependent oligomerization. Fluorescence resonance energy transfer study shows that HspB2 oligomers exchange subunits. Interestingly, HspB2 exhibits target protein-dependent chaperone-like activity: it exhibits significant chaperone-like activity towards dithiothreitol (DTT)-induced aggregation of insulin and heat-induced aggregation of alcohol dehydrogenase, but only partially prevents the heat-induced aggregation of citrate synthase, co-precipitating with the target protein. It also significantly prevents the ordered amyloid fibril formation of a-synuclein. Thus, our study, for the first time, provides biophysical characterization on the structural aspects of HspB2, and shows that it exhibits target protein-dependen

Revisiting Centrioles in Nematodes—Historic Findings and Current Topics

Theodor Boveri is considered as the “father” of centrosome biology. Boveri’s fundamental findings have laid the groundwork for decades of research on centrosomes. Here, we briefly review his early work on centrosomes and his first description of the centriole. Mainly focusing on centriole structure, duplication, and centriole assembly factors in C. elegans, we will highlight the role of this model in studying germ line centrosomes in nematodes. Last but not least, we will point to future directions of the C. elegans centrosome field

Inhibition of Cu<SUP>2+</SUP>- mediated generation of reactive oxygen species by the small heat shock protein &#945;&#946;-crystallin: The relative contributions of the N- and C-terminal domains

Oxidative stress, Cu2+ homeostasis, and small heat shock proteins (sHsp's) have important implications in several neurodegenerative diseases. The ubiquitous sHsp aB-crystallin is an oligomeric protein that binds Cu2+. We have investigated the relative contributions of the N- and C-terminal (C-TD&#945;&#946;-crystallin) domains of &#945;&#946;-crystallin to its Cu2+-binding and redox-attenuation properties and mapped the Cu2+-binding regions. C-TD&#945; &#946;-crystallin binds Cu2+ with slightly less affinity and inhibits Cu2+-catalyzed, ascorbate-mediated generation of ROS to a lesser extent than &#945;&#946; -crystallin. [Cu2+]/[subunit] stoichiometries for redox attenuation by &#945;&#946;-crystallin and C-TD&#945;&#946;-crystallin are 5 and 2, respectively. Both &#945;&#946;-crystallin and C-TD&#945;&#946;-crystallin also inhibit the Fenton reaction of hydroxyl radical formation. Trypsinization of &#945;&#946; -crystallin bound to a Cu2+–NTA column and MALDI-TOF analysis of column-bound peptides yielded three peptides located in the N-terminal domain, and in-solution trypsinization of aB-crystallin followed by Cu2+–NTA column chromatography identified four additional Cu2+-binding peptides located in the C-terminal domain. Thus, Cu2+-binding regions are distributed in the N- and C-terminal domains. Small-angle X-ray scattering and sedimentation-velocity measurements indicate quaternary structural changes in &#945;&#946;-crystallin upon Cu2+ binding. Our study indicates that an oligomer of &#945;&#946;-crystallin can sequester a large number (&#8764; 150) of Cu2+ ions. It acts like a “Cu2+ sponge,” exhibits redox attenuation of Cu2+, and has potential roles in Cu2+ homeostasis and in preventing oxidative stress

Temperature-induced changes in the conformation and chaperone property of HspB2.

<p>(A) far UV-CD spectra of HspB2 at 25°C (1), 45°C (2) and 55°C (3). (B) The change in the mean residue mass ellipticity ([θ]MRM) at 214nm as a function of temperature. (C) Near UV-CD spectra of HspB2 at 25°C (1), 45°C (2) and 55°C (3). (D) The change in the [θ]MRM at 286 nm as a function of temperature. (E) The changes in the fluorescence polarization of AIAS-labelled HspB2 with increasing temperature. (F) % Protection offered by HspB2 to the DTT-induced aggregation of insulin at different temperatures.</p

Far-UV (a) and near-UV (b) CD spectra of HspB2.

<p>Solid line, human HspB2; dotted line, αB-crystallin. The far- and near-UV CD spectra of the proteins, were recorded using 0.1 cm and 1.0 cm path length cuvettes respectively. [θ]MRW is mean residue ellipticity.</p

SDS-PAGE pattern showing purification of HspB2 from the soluble fraction of <i>E.coli</i> BL21(DE3) cells over-expressing human HspB2.

<p>Lane 1, whole cell lysate; Lane 2, soluble fraction; Lane 3, insoluble fraction; respectively, Lane 4, HspB2 precipitated by 20% saturated ammonium sulfate; and Lane 5, HspB2 purified by Phenyl Sepharose chromatography.</p

Chaperone-like activity of human HspB2 towards DTT-induced aggregation of insulin.

<p>(a) Aggregation profile of 0.2 mg/ml insulin alone (1); and in the presence of 1∶0.0625, (2); 1∶0.125, (3); 1∶0.25, (4); and 1∶0.5, (5) ratios of insulin to HspB2 (w/w). (b) The percentage protection of DTT-induced aggregation of insulin by HspB2 as a function of weight ratio of chaperone to target protein.</p

Sedimentation velocity profile of HspB2.

<p>Distribution of sedimentation coefficient of different oligomeric species changes with increasing concentration of HspB2. Panels A, B and C correspond to 0.125, 1 and 3 mg/ml of HSPB2. The numbers depicted above the peaks correspond to molecular masses (in kDa) of the oligomeric species.</p