Native Pr Research Articles

Evaluation of compatibility of native plant growth promoting rhizobacterial isolates against chemical pesticides

Conventional agriculture relies heavily on chemical fertilizers, pesticides and weedicides to control plant diseases, pests and weeds to improve production. Consequently, the soil receives the bulk of complex agrochemical compounds many of which are poisonous to the activity of non-target beneficial soil microorganisms. There is a need to check the tolerance of biocontrol agents like Plant Growth Promoting Rhizobacterial (PGPR) to various pesticides before their practical application. Taking this into account, the investigation was conducted for the screening of the PGPR isolates against chemical pesticide degradation potentialities in Plant Bacteriological Laboratory, Department of Plant Pathology, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur, Nadia, West Bengal during 2021-22. All the native rhizobacterial, Bacillus and Fluorescent pseudomonad isolates tested had good potentiality to tolerate Carbendazim and Glyphosate. Among native Bacillus isolates, BRB89, BRB42, PR 19, PR 20, BRB 52, BRB 56, BRB 74, and SM 9 showed moderate tolerance, whereas among all the native Fluorescent pseudomonads, all except GP2 showed high tolerance against Paraquat. All native Bacillus isolates except PR16 were highly sensitive against Chorpyriphos. However, all the native fluorescent pseudomonads showed tolerance against Chorpyriphos even at 2000 ppm concentration.

Phototransformation of the Red Light Sensor Cyanobacterial Phytochrome 2 from Synechocystis Species Depends on Its Tongue Motifs

Phytochromes are photoreceptors using a bilin tetrapyrrole as chromophore, which switch in canonical phytochromes between red (Pr) and far red (Pfr) light-absorbing states. Cph2 from Synechocystis sp., a noncanonical phytochrome, harbors besides a cyanobacteriochrome domain a second photosensory module, a Pr/Pfr-interconverting GAF-GAF bidomain (SynCph2(1-2)). As in the canonical phytochromes, a unique motif of the second GAF domain, the tongue region, seals the bilin-binding site in the GAF1 domain from solvent access. Time-resolved spectroscopy of the SynCph2(1-2) module shows four intermediates during Pr → Pfr phototransformation and three intermediates during Pfr → Pr back-conversion. A mutation in the tongue's conserved PRXSF motif, S385A, affects the formation of late intermediate R3 and of a Pfr-like state but not the back-conversion to Pr via a lumi-F-like state. In contrast, a mutation in the likewise conserved WXE motif, W389A, changes the photocycle at intermediate R2 and causes an alternative red light-adapted state. Here, back-conversion to Pr proceeds via intermediates differing from SynCph2(1-2). Replacement of this tryptophan that is ∼15 Å distant from the chromophore by another aromatic amino acid, W389F, restores native Pr → Pfr phototransformation. These results indicate large scale conformational changes within the tongue region of GAF2 during the final processes of phototransformation. We propose that in early intermediates only the chromophore and its nearest surroundings are altered, whereas late changes during R2 formation depend on the distant WXE motifs of the tongue region. Ser-385 within the PRXSF motif affects only late intermediate R3, when refolding of the tongue and docking to the GAF1 domain are almost completed.

Abstract 17944: The Impact of Baseline PR Interval on Response to CRT in Heart Failure with Narrow QRS Complexes: A Sub-Study of the ReThinQ Trial

Background: In heart failure (HF) patients (pts) who are treated with cardiac resynchronization therapy (CRT), the programmed AV delay must be shorter than the native PR interval in order to preempt intrinsic, dyssynchronous conduction. When the native PR interval is short, such programming results in truncation of transmitral flow, especially in the presence of dilated atria and marked intraatrial (R–>L) conduction delays. Pts with longer baseline PR intervals can have longer (more physiologic) AV delay programming, thereby allowing complete left atrial emptying. Such pts may show improved response to CRT. We analyzed the impact of baseline PR interval in the ReThinQ Trial, which assessed response to CRT in pts with NYHA class III HF, QRS complex duration <130 ms, and echo-detected dyssynchrony. Methods: Pts randomized to CRT in the ReThinQ trial were divided onto 2 groups depending on baseline PR interval: PR≥180ms and PR<180ms. Outcomes at 6 months were compared in the 2 groups. Results: A total of 87 pts were randomized to CRT; 45 pts had PR≥180ms and 42 pts had PR<180ms. At baseline, age, gender, type of cardiomyopathy, LVEF and QRS duration were similar in the 2 groups. Compared to baseline, peak VO 2 at 6 months (which was the primary endpoint in the ReThinQ Trial) was significantly improved only in pts with PR≥180ms (12.1–>13.5 ml/kg/min, P =0.045) but not in pts with PR<180 ms (12.1–>12.3 ml/kg/min, P =NS). Although both groups had significant improvements in NYHA class, pts with PR≥180ms showed greater improvement at 6 months as compared to pts with PR<180 ms (2.3 vs. 2.6, P =0.017). Finally, improvement in exercise duration showed a trend toward significance in pts with PR≥180ms (9.1–>9.9 min, P =0.12), but there was no difference in pts with PR<180 ms (8.6–>8.7 min, P =0.42). There were no significant changes in LVEF. Conclusions: In the ReThinQ Trial, only pts with PR≥180ms showed significant improvements in peak VO 2 after 6 months of CRT. Similarly, NYHA class improved to a greater degree in pts with PR≥180ms as compared to pts with PR<180 ms. This suggests that some of the benefit of CRT may be negated by truncation of transmitral flow in pts with PR<180 ms. A short baseline PR may be a contributor to CRT non-response.

Time-Resolved WAXS Reveals Accelerated Conformational Changes in Iodoretinal-Substituted Proteorhodopsin

Time-resolved wide-angle x-ray scattering (TR-WAXS) is an emerging biophysical method which probes protein conformational changes with time. Here we present a comparative TR-WAXS study of native green-absorbing proteorhodopsin (pR) from SAR86 and a halogenated derivative for which the retinal chromophore has been replaced with 13-desmethyl-13-iodoretinal (13-I-pR). Transient absorption spectroscopy differences show that the 13-I-pR photocycle is both accelerated and displays more complex kinetics than native pR. TR-WAXS difference data also reveal that protein structural changes rise and decay an order-of-magnitude more rapidly for 13-I-pR than native pR. Despite these differences, the amplitude and nature of the observed helical motions are not significantly affected by the substitution of the retinal's C-20 methyl group with an iodine atom. Molecular dynamics simulations indicate that a significant increase in free energy is associated with the 13-cis conformation of 13-I-pR, consistent with our observation that the transient 13-I-pR conformational state is reached more rapidly. We conclude that although the conformational trajectory is accelerated, the major transient conformation of pR is unaffected by the substitution of an iodinated retinal chromophore.

Biliverdin Amides Reveal Roles for Propionate Side Chains in Bilin Reductase Recognition and in Holophytochrome Assembly and Photoconversion

Linear tetrapyrroles (bilins) perform important antioxidant and light-harvesting functions in cells from bacteria to humans. To explore the role of the propionate moieties in bilin metabolism, we report the semisynthesis of mono- and diamides of biliverdin IXalpha and those of its non-natural XIIIalpha isomer. Initially, these were examined as substrates of two types of NADPH-dependent biliverdin reductase, BVR and BvdR, and of the representative ferredoxin-dependent bilin reductase, phycocyanobilin:ferredoxin oxidoreductase (PcyA). Our studies indicate that the NADPH-dependent biliverdin reductases are less accommodating to amidation of the propionic acid side chains of biliverdin IXalpha than PcyA, which does not require free carboxylic acid side chains to yield its phytobilin product, phycocyanobilin. Bilin amides were also assembled with BV-type and phytobilin-type apophytochromes, demonstrating a role for the 8-propionate in the formation of the spectroscopically native P(r) dark states of these biliprotein photosensors. Neither ionizable propionate side chain proved to be essential to primary photoisomerization for both classes of phytochromes, but an unsubstituted 12-propionate was required for full photointerconversion of phytobilin-type phytochrome Cph1. Taken together, these studies provide insight into the roles of the ionizable propionate side chains in substrate discrimination by two bilin reductase families while further underscoring the mechanistic differences between the photoconversions of BV-type and phytobilin-type phytochromes.

The virus-associated human immunodeficiency virus type 1 Gag-Pol carrying an active protease domain in the matrix region is severely defective both in autoprocessing and in trans processing of gag particles

We have previously demonstrated that a human immunodeficiency virus (HIV) chimeric Gag protein containing a partial replacement of the matrix domain by the viral protease domain (PR) could undergo autoprocessing with no virus particle production [J. Virol. 74 (2000) 3418]. To further analyze the effects of repositioned PR on virus particle production and Gag-Pol incorporation, we introduced the chimeric PR construct into a PR-negative Gag-Pol expression plasmid and coexpressed the resultant construct with a Pr55 gag expression plasmid (pGAG) in 293T cells. Analysis indicated that the chimeric PR was similar to native PR in that both could prevent virus particle production in cotransfections with an equivalent amount of pGAG plasmid DNA, suggesting an efficient trans processing of Pr55 gag by the chimeric PR. In cotransfections with the pGAG at a DNA ratio of 1:10 to 1:20, which resembles the normal intracellular expression ratio of Gag-Pol to Gag, Gag-Pol carrying the PR in the Gag coding region could undergo autoprocessing in cells and was incorporated into virus particles at a level about 20–40% of that of wild-type Gag-Pol. However, the incorporated chimeric Gag-Pol was unable to autocleave and unable to process the Gag particles properly, as mature particle-associated reverse transcriptase (RT) and p24 gag proteins were barely detected. Our data strongly suggest that positioning an active HIV PR in the matrix region significantly affects the PR-mediated virus particle maturation.

Cardiomyocytes Bind and Activate Renal But not Non-Renal Human Prorenin.

P10 Cardiomyocytes bind and internalize recombinant human prorenin (rhPR) and renin via mannose 6-phosphate (M6P) receptors (van Kesteren et al., Hypertension 1997). PR binding is followed by intracellular activation. Here we investigated 1) whether cardiomyocytes bind native human (pro)renin ((P)R) from various sources and 2) whether PR binding is followed by activation. Myocytes were isolated from Wistar rat pups by enzymatic dissociation and grown to confluence (6 x 10 5 cells in 3.8 cm 2 wells). Cells were incubated with rhPR, human plasma (obtained from captopril-treated patients (CAP), patients with renal artery stenosis (RAS) and anephric subjects (Nx)), or human amniotic fluid (HAF), diluted in DMEM (final volume 0.4 mL), for 4 hrs at 37°C, either with or without 10 mM M6P. Cell-associated R and PR, the latter following prior activation by plasmin, were determined by enzyme-kinetic assay. Results: See Table. Intracellular PR activation is reflected by the higher R/PR ratio in the cell lysates as compared to the medium. Conclusions: Cardiomyocytes bind and activate native human PR from renal (CAP & RAS), but not from non-renal (Nx & HAF) sources in a M6P- dependent manner. These data suggest that renal PR is meant to be sequestered by nonrenin-producing tissues in order to contribute to local angiotensin production, whereas non-renal PR functions at its production site only.

Cardiomyocytes Bind and Activate Renal But not Non-Renal Human Prorenin.

P10 Cardiomyocytes bind and internalize recombinant human prorenin (rhPR) and renin via mannose 6-phosphate (M6P) receptors (van Kesteren et al., Hypertension 1997). PR binding is followed by intracellular activation. Here we investigated 1) whether cardiomyocytes bind native human (pro)renin ((P)R) from various sources and 2) whether PR binding is followed by activation. Myocytes were isolated from Wistar rat pups by enzymatic dissociation and grown to confluence (6 x 10 5 cells in 3.8 cm 2 wells). Cells were incubated with rhPR, human plasma (obtained from captopril-treated patients (CAP), patients with renal artery stenosis (RAS) and anephric subjects (Nx)), or human amniotic fluid (HAF), diluted in DMEM (final volume 0.4 mL), for 4 hrs at 37°C, either with or without 10 mM M6P. Cell-associated R and PR, the latter following prior activation by plasmin, were determined by enzyme-kinetic assay. Results: See Table. Intracellular PR activation is reflected by the higher R/PR ratio in the cell lysates as compared to the medium. Conclusions: Cardiomyocytes bind and activate native human PR from renal (CAP & RAS), but not from non-renal (Nx & HAF) sources in a M6P- dependent manner. These data suggest that renal PR is meant to be sequestered by nonrenin-producing tissues in order to contribute to local angiotensin production, whereas non-renal PR functions at its production site only.

Pigment of the imagination: a history of phytochrome research

Daylength and flowering the 1930s spectral studies action spectra for floral induction action spectra for elongation growth photoreversibility the 1950s high-energy responses detection partial purification the properties of phytochrome phototransformation in vivo spectrophotometric studies seed germination and flowering photoperiodic timing mougeotia ode of action the Hartmann model large phytochrome the gene regulation hypothesis dark transformations localization properties of large phytochrome the membrane hypothesis high-irradiance responses the very low fluence response - a dimer model shade avoidance the chromophore native phytochrome native pr and pfr sequestered and ubiquitinated phytochrome multiple phytochromes gene regulation autoregulation phytochrome and flowering. Epiloque: weed control coloured mulch.

Surface enhanced resonance Raman scattering (SERRS) as a probe of the structural differences between the Pr and Pfr forms of phytochrome

Surface enhanced resonance Raman scattering (SERRS) spectra have been obtained from the active, far-red light absorbing (Pfr) and biologically inactive (Pr) forms of phytochrome adsorbed on silver colloids. Substantial differences between the SERRS spectra of the two forms in the low and high wavenumber regions are observed using 406.7 nm wavelength excitation. These differences reinforce those seen with 413.1 nm wavelength excitation in the high wavenumber region. Simultaneously, extensive differences are observed in the SERRS obtained from the same form in the low wavenumber region using 406.7 nm, as compared with 413.1 nm wavelength excitation. The relative intensity differences observed for the two forms, and those obtained using two slightly different excitation wavelengths to illuminate the same form, suggest that some type of subtle, proteincontrolled structural variation is responsible for the spectroscopic differences. A Z → E isomerization during the Pr → Pfr phototransformation is consistent with the SERRS data, although the overall chromophore conformations are most likely conserved for the native Pr- and Pfr-phytochrome species. Slight out-of-plane ring twisting, accompanying the Pr → Pfr photoisomerization, may be responsible for the large difference in the spectroscopic properties of the native Pr and Pfr chromophores.

THE KINETICS OF THE EARLY STAGES OF THE PHYTOCHROME PHOTOTRANSFORMATION Pr→ Pfr. A COMPARATIVE STUDY OF SMALL (60 kDalton) and NATIVE (124 kDalton) PHYTOCHROMES FROM OAT

Abstract—A comparative study of the decay kinetics of photogenerated transients from small (60 kDalton) and native (124 kDalton) oat phytochrome in the red‐absorbing form (Pr) in phosphate buffer containing 5 mM ethylenediamine tetraacetic acid, pH 7.8, (PB) and in PB containing 20% ethylene glycol, has been carried out in the temperature range 275–298 K. The analysis confirmed that at least two primary photoproducts, intermediates Ii700s and Ii7oo are formed from Pr. The kinetic parameters, as observed in PB at 695 nm and 275 K, are similar for the I700 intermediates of both small and native phytochrome. Namely, the lifetimes are about 21 μs (component percentages 38%) for the I Ii700s and about 200 μ.s (62%) for the Ii700S‐ Arrhenius preexponential factors (A) of about 1016 and 1015 s‐1and activation energies of about 61 and 56 kJ/mol were measured for the absorbance decays of the I700S of small and native phytochrome, respectively. The kinetic data favour parallel paths for the formation of the Ii700s from Pr, and the activation parameters indicate that the primary photoreactions of the transformation from Pr to the far‐red‐absorbing form are restricted to the chromophore within the protein. Moreover, the relatively modest temperature dependence of the lifetimes of the Ii700S from small and native Pr supports the working hypothesis that the ground state reactions to the Iibl, intermediates–although somewhat influenced by the polypeptide fragment that is removed upon degradation of native to small Pr–are localized to the chromophore, as is most probably the case also for the primary photoreactions. The effect of the addition of 20% ethylene glycol on the pre‐exponential factors of the time‐dependent decay functions is discussed in similar terms of the early stages of the phototransformation.

Chromophore Content and Molar Absorptivity of Phytochrome in the Pr Form

Abstract Quantitative absorption spectra in the visible and UV region were recorded for denaturated phytochrome (P r) and phytochromobilin peptides in comparison with native Pr. The method was tested with C-phycocyanin from Spirulina platensis. Based on known molar absorptivities for denaturated phycocyanin and suitable model compounds, and on the ratio A620 native/A655-665 degraded = 2.9, ^native was determined to be 102000 m-1 cm-1 for one phycocyanobilin chromophore in native phycocyanin. Likewise, Native was calculated to be 19000 M-1 cm-1. The corresponding ratios for Pr were Anative/Adenatured = 3.4 and Anative/Adegraded = 3.7, this yielded ε665 native = 109000 to 118000 and ε380 native = 36000 M-1 cm-1 for phytochromobilin in native P r. This value corresponds to one phytochromobilin per small phytochrome (60000 D) if data from the literature are corrected for the content of colorless proteins. The latter has been asessed from (i) the purity index (A280/A665) and (ii) the contribution of the phytochromobilin chromophore at 280 nm as derived from model compounds.