Studies on oxygen evolution of inside-out thylakoid vesicles from mangroves: Chloride requirement, pH dependence and polypeptide composition

Close similarity between the rat native low-conductance K+ channel in the apical membrane of renal cortical collecting duct principal cells and the cloned rat ROMK channel strongly suggest that the two are identical. Prominent features of ROMK regulation are a steep pH dependence and activation by protein kinase A (PKA)-dependent phosphorylation. In this study, we investigated the pH dependence of cloned renal K+ channel (ROMK2), wild-type (R2-WT), and PKA site mutant channels (R2-S25A, R2-S200A, and R2-S294A). Ba2+-sensitive outward whole cell currents (holding voltage −50 mV) were measured in two-electrode voltage-clamp experiments in Xenopus laevisoocytes expressing either R2-WT or mutant channels. Intracellular pH (pHi) was measured with pH-sensitive microelectrodes in a different group of oocytes from the same batch on the same day. Resting pHi of R2-WT and PKA site mutants was the same: 7.32 ± 0.02 ( n = 22). The oocytes were acidified by adding 3 mM Na butyrate with external pH (pHo) adjusted to 7.4, 6.9, 6.4, or 5.4. At pHo 7.4, butyrate led to a rapid (τ: 163 ± 14 s, where τ means time constant, n= 4) and stable acidification of the oocytes (ΔpHi0.13 ± 0.02 pH units, where Δ means change, n = 12). Intracellular acidification reversibly inhibited ROMK2-dependent whole cell current. The effective acidic dissociation constant (p K a) value of R2-WT was 6.92 ± 0.03 ( n = 8). Similarly, the effective p K a value of the N-terminal PKA site mutant R2-S25A was 6.99 ± 0.02 ( n = 6). The effective p K a values of the two COOH-terminal PKA site mutant channels, however, were significantly shifted to alkaline values; i.e., 7.15 ± 0.06 ( n = 5) for R2-S200A and 7.16 ± 0.03 ( n = 8) for R2-S294A. The apparent ΔpH shift between the R2-WT and the R2-S294A mutant was 0.24 pH units. In excised inside-out patches, alkaline pH 8.5 activated R2-S294A channel current by 32 ± 6.7%, whereas in R2-WT channel patches alkalinzation only marginally increased current by 6.5 ± 1% ( n = 5). These results suggest that channel phosphorylation may substantially influence the pH sensitivity of ROMK2 channel. Our data are consistent with the hypothesis that in the native channel PKA activation involves a shift of the pKa value of ROMK channels to more acidic values, thus relieving a H+-mediated inhibition of ROMK channels.

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EDTA-induced release of manganese and proteins from inside-out thylakoid vesicles and the inhibition of oxygen evolution

Biochemical and Biophysical Research Communications ◽

10.1016/0006-291x(83)91184-1 ◽

1983 ◽

Vol 110 (2) ◽

pp. 545-551 ◽

Cited By ~ 10

Author(s):

R.W. Mansfield ◽

J. Barber

Keyword(s):

Oxygen Evolution ◽

Inside Out

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Biochemical characteristics of myofibrillar proteins of the fish parasite Isoparorchis hypselobagri (Trematoda) as survival traits in an oxygen-rich environment

Biological Letters ◽

10.2478/v10120-012-0009-0 ◽

2012 ◽

Vol 49 (1) ◽

pp. 45-58 ◽

Cited By ~ 1

Author(s):

Usma Abbasi ◽

Riaz Ahmad ◽

Absar-ul Hasnain

Keyword(s):

Atpase Activity ◽

Striated Muscle ◽

Ph Dependence ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Inactivation Kinetics ◽

Myofibrillar Proteins ◽

Swim Bladder ◽

Polypeptide Composition ◽

Wallago Attu

Abstract We have investigated biochemical properties of myofibrillar proteins of the digenetic trematode Isoparorchis hypselobagri, which correlate with its survival in the oxygen-rich swim bladder of its host catfish (Wallago attu). The polypeptide composition of the trematode’s natural actomyosin (NAM) was striated-muscle-like, with the exception that a 98-kD polypeptide corresponding to paramyosin also existed in its sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) profiles. The profiles of immunoprecipitated NAM of the trematode support these inferences. Ca2+-sensitivity of myofibrillar contractility and Mg2+-ATPase activity of I. hypselobagri resembled troponin-linked calcium regulation of the host striated muscle. Myofibrillar permeability to water influx was insensitive to calcium chelation at neutral pH. However, the host swim bladder myofibrils displayed smooth-muscle-like polypeptide composition, pH dependence of contractility, Ca2+-sensitivity, ATPase activities, and inactivation kinetics. We propose 2 survival strategies that I. hypselobagri appears to have co-evolved: (i) fast-muscle-like musculature with exceptionally high contractility or ATPase activity; and (ii) type-II myosin resembling the host muscle in functional plasticity.

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A Study of the Anomalous pH Dependence of the Oxygen Evolution Reaction at Platinum Electrodes in Acid Solutions

Journal of The Electrochemical Society ◽

10.1149/1.2120064 ◽

1983 ◽

Vol 130 (8) ◽

pp. 1694-1699 ◽

Cited By ~ 20

Author(s):

V. I. Birss ◽

A. Damjanovic

Keyword(s):

Oxygen Evolution ◽

Oxygen Evolution Reaction ◽

Ph Dependence ◽

Platinum Electrodes ◽

Acid Solutions

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Effect of α-, β-and γ-Cyclodextrins on Oxygen Evolution by the Thylakoid Membrane. Influence of pH and Temperature

Zeitschrift für Naturforschung C ◽

10.1515/znc-2001-9-1018 ◽

2001 ◽

Vol 56 (9-10) ◽

pp. 792-802 ◽

Cited By ~ 2

Author(s):

Govindachary Sridharan ◽

Simon Gaudreau ◽

Laetitia Dalstein ◽

Christelle Huiban ◽

Agnès Lejeune ◽

...

Keyword(s):

Oxygen Evolution ◽

Thylakoid Membrane ◽

Ph Dependence ◽

Thylakoid Membranes ◽

Red Shift ◽

Chemical Effect ◽

Second Derivative ◽

Important Conclusion ◽

Derivative Spectra ◽

Second Derivative Spectra

AbstractThe present work investigates the effect of α-, β- and γ-cyclodextrins (CD), i.e., α-CD, β- CD and γ-CD, on the oxygen evolution activity, the protein content and the uv-vis spectroscopic characteristics of thylakoid membranes. To study the pH-dependence, the thylakoids were incubated with the cyclodextrins at 273 K for a period of 10 min in the pH range from 5.5 to 9.0. To study the temperature-dependence the membranes were incubated at 273 and 293 K at pH 6.5, that is, the pH which induces a maximal oxygen evolution in the thylakoid preparations. The major observations are: (i) a stimulation of oxygen evolution in thylakoids incubated with α- and β-CD either in acidic or alkaline conditions, (ii) a low inhibitory effect induced by γ-CD on oxygen evolution, (iii) a significant decrease of the stimulatory effect of α- and β-CD on oxygen evolution as the incubation temperature is raised from 273 to 293 K, (iv) the apparent inability of the cyclodextrins to change the protein contents of the thylakoids, and (v) a significant CD-induced red-shift from 681 to 683 nm observed in the absorption and second derivative spectra of the thylakoid membranes treated with β-CD. First, it was found that the temperature effect described here is in accord with the general trend of the chemical effect of various cyclodextrins, i.e., the increase of the CD efficiency with decreasing temperature. Secondly, the CD effect is related to the size of the inner cavity diameter of the cyclodextrin molecules. An important conclusion in this work is that the molecular targets of the cyclodextrins are not limited to the thylakoid lipids as was described previously [Rawyler A. and Siegenthaler P.A. (1996) Biochim. Biophys. Acta 1278, 89-97], but are located as well in other molecular species exposed at the stromal side of the thylakoid membrane. In particular, the CD-induced red-shift from 681 to 683 nm in the absorption and second derivative spectra of the thylakoid membranes indicates that the cyclodextrins targets might be either the exposed heme macrocycle in cytochrome b559, or the chlorophylls and pheophytins in the pigment-proteins of the photosystems I and II.

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