Functional characterization of a minimal potassium channel expressed from a synthetic gene

Hausdorff, Sharon F.; Goldstein, Steve A.N.; Rushin, Elizabeth E.; Miller, Christopher

doi:10.1021/bi00227a025

Cited by 56 publications

(28 citation statements)

References 39 publications

(56 reference statements)

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“…The lack of an apparent voltage sensor and the small size of IsK with a single membrane spanning domain created skepticism of its potassium channel function when its clone first appeared (Hausdorff, Goldstein, Rushin, and Miller, 1991;Goldstein and Miller, 1991). The study of mutations of this channel even revealed that only part of this protein was sufficient to generate its potassium channel activity (Takumi, Moriyoshi, Aramori, Ishii, Oiki, Okada, Ohkubo, and Nakanishi, 1991).…”

Section: Discussionmentioning

confidence: 99%

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Cui

Kline

Pennefather

et al. 1994

The Journal of General Physiology

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ABSTRACTlsK is a K + channel of the delayed rectifier type widely distributed throughout both excitable and nonexcitable cells. Its structure is different from other cloned K § channels and molecular details of its gating remain obscure. Here we show that the activation kinetics of IsK expressed in Xenopus oocytes depend upon the amount of its mRNA injected, with larger amounts resulting in slower activation kinetics with a longer initial delay during activation. Similar changes in activation kinetics occur with time after a single injection of IsK mRNA. We present two kinetic schemes which illustrate how our experimental results could arise. Both imply an interaction among individual channel proteins during IsK activation. The dependence of channel gating on mRNA concentration provides a novel mechanism for long term regulation of ion current kinetics.

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Section: Discussionmentioning

confidence: 99%

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Cui

Kline

Pennefather

et al. 1994

The Journal of General Physiology

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“…It should be noted that despite the remarkably diverse array of membranes ( e g from bacteria, plants, and animals) that have retained this channel pore sequence in K+ channels, there is a nove1 exception to this molecular basis for selective K+ conductance. A small (130 amino acids, approximately 15 kD) protein (the "mink" or minimal K+ channel) has recently been identified in mammalian kidney, uterus, and heart that is structurally unrelated to the ubiquitous, large family of 70-kD K+ channels that share the conserved selectivity filter (Hausdorff et al, 1991).…”

Section: Lmmunodetection Of K+ Channel Proteins In the Lnner Envelopementioning

confidence: 99%

Characterization of a Chloroplast Inner Envelope K+ Channel

Peters

Berkowitz

1994

Plant Physiol.

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A K+-conducting protein of the chloroplast inner envelope was characterized as a K+ channel. Studies of this transport protein in the native membrane documented its sensitivity to K+ channel blockers. Further studies of native membranes demonstrated a sensitivity of K+ conductance to divalent cations such as MgZ+, which modulate ion conduction through interaction with negative surface charges on the inner-envelope membrane. Purified chloroplast inner-envelope vesicles were fused into an artificial planar lipid bilayer to facilitate recording of single-channel K+ currents. These single-channel K+ currents had a slope conductance of 160 picosiemens. Antibodies generated against the conserved amino acid sequence that serves as a selectivity filter in the pore of K+ channels immunoreacted with a 62-kD polypeptide derived from the chloroplast inner envelope. This polypeptide was fractionated using density gradient centrifugation. Comigration of this immunoreactive polypeptide and K+ channel activity in sucrose density gradients further suggested that this polypeptide is the protein facilitating K+ conductance across the chloroplast inner envelope.Studies with intact chloroplasts (Wu and Berkowitz, 1992) indicate that K+ flux across the inner envelope occurs through a well-regulated transport pathway that is likely a K+-conducting ion channel. K' flux into or out of the chloroplast stroma is indirectly linked to H+ counterexchange (Maury et al., 1981; Wu and Berkowitz, 1992). Movement of K+ through this channel protein, therefore, has a profound effect on stromal pH and, hence, photosynthesis (Werdan et al., 1975;Maury et al., 1981; Wu and Berkowitz, 1992). Previous work from this laboratory has demonstrated that a K'-conducting protein can be successfully detergent-solubilized from preparations of purified inner-envelope membrane vesicles and functionally reconstituted into artificial liposomes (Wang et al., 1993). This putative K+ channel protein has not previously been characterized. In the work reported here, we used severa1 approaches to characterize the nature of this K+ transport protein. Results of these experiments are consistent with the presence of a K+ channel in the inner envelope that shares some structural, functional, and regulatory properties with K' channels from other membrane systems ( e g Rudy et al., 1991; Anderson et al., 1992). 955 MATERIALS AND METHODS Preparation of Chloroplast Inner-Envelope Membrane VesiclesInner-envelope vesicles were prepared from spinach (Spinacia oleracea L.) as described previously (Berkowitz and Peters, 1993). Briefly, intact chloroplasts (isolated from 5-10 kg of spinach leaves using Perco11 step gradients) were exposed to freeze/thaw cycles in hyperosmotic medium, and the thylakoids were removed before the crude membrane fraction was loaded on a discontinuous SUC step gradient. Inner-envelope vesicles were removed from the 0.8 ~/ 0 . 4 6 M Suc interface of the gradient, washed in envelope medium (0.2 M SUC, 2 mM Na2EDTA, 2 mM DTT, and 10 m~ TricineNaOH, pH 7.5), a...

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“…Members of the second K ÷ channel type have been termed minK channels [2]. This type of channel has been originally cloned from rat kidney and was subsequently found in heart and uterus tissue of different species [3][4][5][6].…”

Section: Introductionmentioning

confidence: 99%

Effects of [Ca²⁺]_i and temperature on minK channels expressed in Xenopus oocytes

Büsch¹,

Läng²

1993

FEBS Letters

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Slowly activating, voltage-dependent minK channels cloned from rat kidney were expressed in Xenopus oocytes. Increase in the bath temperature from 22 to 32°C resulted in a dramatic acceleration of minK channel activation. The extraordinarily high Q~0 of minK channel activation was voltage-dependent, being higher at more negative potentials (Q10 at -20 mV: 7.02; at 20 mV: 4.0). While activation of minK channels was highly voltage-dependent at 22°C, voltage had only little effect on minK channel activation at 32°C. Increase in [Ca2+]~, which has recently been shown to increase the maximal conductance gmax at room temperature, did not affect g .... at 32°C. However, increase of [CaZ+]i caused acceleration of minK channel activation at both temperatures. The interaction of [Ca2+]i and temperature on gmax and activation rate of minK channels described here is very similar to recent findings on Ca-and temperature-effects on the slowly activating potassium conductance/Ks in guinea pig cardiac myocytes.

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Functional characterization of a minimal potassium channel expressed from a synthetic gene

Cited by 56 publications

References 39 publications

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Characterization of a Chloroplast Inner Envelope K+ Channel

Effects of [Ca²⁺]_i and temperature on minK channels expressed in Xenopus oocytes

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Functional characterization of a minimal potassium channel expressed from a synthetic gene

Cited by 56 publications

References 39 publications

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Gating of IsK expressed in Xenopus oocytes depends on the amount of mRNA injected.

Characterization of a Chloroplast Inner Envelope K+ Channel

Effects of [Ca2+]i and temperature on minK channels expressed in Xenopus oocytes

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Effects of [Ca²⁺]_i and temperature on minK channels expressed in Xenopus oocytes