Modulation of Voltage-dependent Shaker Family Potassium Channels by an Aldo-Keto Reductase*

Abstract

The β subunit (Kvβ) of the Shaker family voltage-dependent potassium channels (Kv1) is a cytosolic protein that forms a permanent complex with the channel. Sequence and structural conservation indicates that Kvβ resembles an aldo-keto reductase (AKR), an enzyme that catalyzes a redox reaction using an NADPH cofactor. A putative AKR in complex with a Kv channel has led to the hypothesis that intracellular redox potential may dynamically influence the excitability of a cell through Kvβ. Since the AKR function of Kvβ has never been demonstrated, a direct functional coupling between the two has not been established. We report here the identification of Kvβ substrates and the demonstration that Kvβ is a functional AKR. We have also found that channel function is modulated when the Kvβ-bound NADPH is oxidized. Further studies of the enzymatic properties of Kvβ seem to favor the role of Kvβ as a redox sensor. These results suggest that Kvβ may couple the excitability of the cell to its metabolic state and present a new avenue of research that may lead to understanding of the physiological functions of Kvβ. The β subunit (Kvβ) of the Shaker family voltage-dependent potassium channels (Kv1) is a cytosolic protein that forms a permanent complex with the channel. Sequence and structural conservation indicates that Kvβ resembles an aldo-keto reductase (AKR), an enzyme that catalyzes a redox reaction using an NADPH cofactor. A putative AKR in complex with a Kv channel has led to the hypothesis that intracellular redox potential may dynamically influence the excitability of a cell through Kvβ. Since the AKR function of Kvβ has never been demonstrated, a direct functional coupling between the two has not been established. We report here the identification of Kvβ substrates and the demonstration that Kvβ is a functional AKR. We have also found that channel function is modulated when the Kvβ-bound NADPH is oxidized. Further studies of the enzymatic properties of Kvβ seem to favor the role of Kvβ as a redox sensor. These results suggest that Kvβ may couple the excitability of the cell to its metabolic state and present a new avenue of research that may lead to understanding of the physiological functions of Kvβ. Kv channels control the flow of K+ through the cell membrane in response to changes in membrane potential. Opening of the channel causes membrane hyperpolarization that can curtail excessive membrane excitability or simply tone down normal membrane activity. found in and to the cytosolic of the channels to a complex Kvβ and have a with to an AKR aldo-keto aldo-keto The of the of a that has a AKR with in the to and has a These suggest that Kvβ is a functional AKR. changes in Kv channel have been when the redox state is or the Kvβ the between redox and Kv channel the of Kvβ is a functional AKR and the AKR function is to channel the of an to an an NADPH cofactor. have a that in to also as The a in an NADPH and an to a The redox reaction when the enzyme a the to the The an the enzyme and is the to NADPH its The is and the or the protein changes with and The in is that an can to an the of an to and protein a between the and with a or a in using the and through the and to in to an of and with the to and protein a the the and protein using to the and protein with the a with a to of The protein a The with the reaction using the the reaction the reaction and of substrates in the reaction The reaction and NADPH using a with a The reaction the of a The the of the in the with protein and the NADPH a protein of and its using the in potassium and We the reaction the enzyme and the We a of of of to the of in the reaction protein to and with of and the The of the the NADPH is between the and the studies have that the is to the NADPH The of NADPH Kvβ and the with a function in The of the is as the in the or to the and in and using the channel and a of The has a that can to channel and a or of that the in to the in also and with in the of and the results and to a of The and The and with The in the is to the or K+ the and to The of the is and of of new flow with a flow of of with or has been can an function with two The is also the is the and the is to to the the to channel with the the and modulated Since the and not the in the is to a direct the or a of in the We the and the of the as the of channel and the as a the of Kvβ channels with in a new in or and to the The of or is in NADPH as a Since potassium channels the intracellular the to potassium protein and the and protein the of the as a in the and the of NADPH is Since has the the the as a AKR We have potential Kvβ substrates and two of the and here to the enzymatic of Kvβ and channel properties of a of The is in the the protein with The of and in the of the the and and the The functions to the of and and with and The of to is has of the the Kvβ with of the with The of is to the in the protein with The and the of the the of the Since NADPH is a the of NADPH the a function with a of with a results when with a of that the the that the NADPH is to an enzymatic control NADPH with in the of and the in NADPH A of NADPH a of that the reaction is when the to the an to an and the protein and is in and a the of in as reductase and and The protein has a NADPH as an the protein with the of the with a of also The is that of the that the down the enzymatic the that Kvβ is a functional studies of of the reaction or of the reaction or the that the of the is of to the the and the The of the with a of to the that of a an to an the protein its NADPH the reaction through a found that the not a to indicates that the to the to the and found potential The of is in with the and to the as The of NADPH with a function with a of with a The and the of the NADPH and that Kvβ is a functional the the of the NADPH the or We found that the with or and not of a a the protein and its The is that of the and the that that Kvβ is a functional is that the not with indicates that substrates have the enzyme that is not that is not indicates that Kvβ is of the NADPH a to or a that is in of redox in a cell is a in the enzymatic reaction the the in The or and the a with a of and and a of and and NADPH with not that Kvβ has to NADPH The and is the is the is with a reaction in the Since the not that the is when Kvβ catalyzes its may have physiological an control and to the that Kvβ is an to a A in the We and the and the the that of the The is that of NADPH the of a Kvβ indicates that the is and the that Kvβ is a functional Kvβ Kvβ is a functional AKR to the of the redox reaction Kvβ channel with and channel function and a Kvβ a an and the intracellular of the channel to A and and of and the and the an response not We have the of channel the and the the substrates Kvβ of with with the with the and with the two The the in the the to and the channel and The of and the in A and the the to the and the in and the and to and to the of channel to and and of and the that the substrates channel function through the in channel function is to the redox reaction with the or the the of with the the changes to and and of the of with the changes to and and of a in channel and the of that is in channel when the Kvβ is the studies that the substrates channel functions through and through the redox of the NADPH a Kvβ channel an Kvβ an and when a is to the the NADPH is oxidized. The a in channel hypothesis is can the two and that a with NADPH channel the with the cofactor. the when the with of NADPH of in the and in two of the to the in channel when NADPH to These suggest that Kvβ an can an enzymatic when to its and NADPH to the in channel function as a found that when a and to a channel and a state that not the a in channel can the of channel the as a function of a Since with the NADPH a that channel may also the is channel channel using and in the of the The functions with a of and and channel function to the in the the that channel is the Kvβ a of with the The indicates the the the channel a of and The functions to the of and Kvβ with the studies the reaction that the of channel can with that of NADPH in a We found that and the NADPH with a of and that in the with the of channel in is that the is a Kvβ is with a Kvβ in an may have properties that in have that Kvβ is a functional and have that Kvβ substrates channel through the NADPH cofactor. These results that the functions of the two dynamically is that of Kvβ with channels channel and properties of channel and the demonstration that Kvβ is a functional is to of channel functions to the of the NADPH as a the the and AKR the is in of the AKR and in Kvβ is an a when the to a has the the a of the to an in an reductase the a of the of the substrates These results that the may an role in and the in Kvβ the of its enzymatic of is and is the the two catalyzes a a of the the of Kvβ is with a the the NADPH a of is Kvβ. These results that Kvβ is a enzyme has a Kvβ a redox that in and channel function or an enzyme is channel here between the two seem to favor the redox The enzymatic the function of Kvβ is to not is to redox the and the with structural studies have that the NADPH a of the the present in the of The is to a the NADPH and the of the in in The of Kvβ that the complex is to to a redox and the channel in a modulated state a of with Kvβ has been in channel and and have found that a the Kvβ-bound NADPH and also channel functions in a not a to the understanding the of the functional coupling and the physiological of functional between the two a of coupling intracellular redox to the excitability of a The results here to understanding the physiological functions of Kvβ. Kv channels control the flow of K+ through the cell membrane in response to changes in membrane potential. Opening of the channel causes membrane hyperpolarization that can curtail excessive membrane excitability or simply tone down normal membrane activity. found in and to the cytosolic of the channels to a complex Kvβ and have a with to an AKR aldo-keto aldo-keto The of the of a that has a AKR with in the to and has a These suggest that Kvβ is a functional AKR. changes in Kv channel have been when the redox state is or the Kvβ the between redox and Kv channel the of Kvβ is a functional AKR and the AKR function is to channel the of an to an an NADPH cofactor. have a that in to also as The a in an NADPH and an to a The redox reaction when the enzyme a the to the The an the enzyme and is the to NADPH its The is and the or the protein changes with and The in is that an can to an the of an to and protein a between the and with a or a in using the and through the and to in to an of and with the to and protein a the the and protein using to the and protein with the a with a to of The protein a 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the of channel and the as a the of Kvβ channels with in a new in or and to the The of or is in NADPH as a Since potassium channels the intracellular the to potassium protein and the and and protein a between the and with a or a in using the and through the and to in to an of and with the to and protein a the the and protein using to the and protein with the a with a to of The protein a The with the reaction using the the reaction the reaction and of substrates in the reaction The reaction and NADPH using a with a The reaction the of a The the of the in the with protein and the NADPH a protein of and its using the in potassium and We the reaction the enzyme and the We a of of of to the of in the reaction protein to and with of and the The of the the NADPH is between the and the studies have that the is to the NADPH The of NADPH Kvβ and the with a function in The of the is as the in the or to the and in and using the channel and a of The has a that can to channel and a or of that the in to the in also and with in the of and the results We and to a of The and The and with The in the is to the or K+ the and to The of the is and of of new flow with a flow of of with or has been can an function with two The is also the is the and the is to to the the to channel with the the and modulated Since the and not the in the is to a direct the or a of in the We the and the of the as the of channel and the as a the of Kvβ in or and to the The of or is in NADPH as a Since potassium channels the intracellular the to potassium protein and the and protein the of the as a in the and the of NADPH is Since has the the the as a AKR We have potential Kvβ substrates and two of the and here to the enzymatic of Kvβ and channel in the protein with The and the of the the of the Since NADPH is a the of NADPH the a function with a of with a results when with a of that the the that the NADPH is to an enzymatic control NADPH with in the of and the in NADPH A of NADPH a of that the reaction is when the to the an to an and the protein and is in and a the of in as reductase and and The protein has a NADPH as an the protein with the of the with a of also The is that of the that the down the enzymatic the that Kvβ is a functional studies of of the reaction or of the reaction or the that the of the is of to the the and the The of the with a of to the that of a an to an the protein its NADPH the reaction through a found that the not a to indicates that the to the to the and found potential The of is in with the and to the as The of NADPH with a function with a of with a The and the of the NADPH and that Kvβ is a functional the the of the NADPH the or We found that the with or and not of a a the protein and its The is that of the and the that that Kvβ is a functional is that the not with indicates that substrates have the enzyme that is not that is not indicates that Kvβ is of the NADPH a to or a that is in of redox in a cell is a in the enzymatic reaction the the in The or and the a with a of and and a of and and NADPH with not that Kvβ has to NADPH The and is the is the is with a reaction in the Since the not that the is when Kvβ catalyzes its may have physiological an control and to the that Kvβ is an to a A in the We and the and the the that of the The is that of NADPH the of a Kvβ indicates that the is and the that Kvβ is a functional Kvβ Kvβ is a functional AKR to the of the redox reaction Kvβ channel with and channel function and a Kvβ a an and the intracellular of the channel to A and and of and the and the an response not We have the of channel the and the the substrates Kvβ of with with the with the and with the two The the in the the to and the channel and The of and the in A and the the to the and the in and the and to and to the of channel to and and of and the that the substrates channel function through the in channel function is to the redox reaction with the or the the of with the the changes to and and of the of with the changes to and and of a in channel and the of that is in channel when the Kvβ is the studies that the substrates channel functions through and through the redox of the NADPH a Kvβ channel an Kvβ an and when a is to the the NADPH is oxidized. The a in channel hypothesis is can the two and that a with NADPH channel the with the cofactor. the when the with of NADPH of in the and in two of the to the in channel when NADPH to These suggest that Kvβ an can an enzymatic when to its and NADPH to the in channel function as a found that when a and to a channel and a state that not the a in channel can the of channel the as a function of a Since with the NADPH a that channel may also the is channel channel using and in the of the The functions with a of and and channel function to the in the the that channel is the Kvβ a of with the The indicates the the the channel a of and The functions to the of and Kvβ with the studies the reaction that the of channel can with that of NADPH in a We found that and the NADPH with a of and that in the with the of channel in is that the is a Kvβ is with a Kvβ in an may have properties that in protein the of the as a in the and the of NADPH is Since has the the the as a AKR We have potential Kvβ substrates and two of the and here to the enzymatic of Kvβ and channel in the protein with The and the of the the of the Since NADPH is a the of NADPH the a function with a of with a results when with a of that the the that the NADPH is to an enzymatic control NADPH with in the of and the in NADPH A of NADPH a of that the reaction is when the to the an to an and the protein and is in and a the of in as reductase and and The protein has a NADPH as an the protein with the of the with a of also The is that of the that the down the enzymatic the that Kvβ is a functional AKR. the that of a an to an the protein its NADPH the reaction through a found that the not a to indicates that the to the to the and found potential The of is in with the and to the as The of NADPH with a function with a of with a The and the of the NADPH and that Kvβ is a functional AKR. the the of the NADPH the or We found that the with or and not of a a the protein and its The is that of the and the that that Kvβ is a functional AKR. is that the not with indicates that substrates have the enzyme that is not that is not indicates that Kvβ is of the NADPH a to or a that is in of redox in a cell The is a in the enzymatic reaction the the in The or and the a with a of and and a of and and NADPH with not that Kvβ has to NADPH The and is the is the is with a reaction in the Since the not that the is when Kvβ catalyzes its may have physiological an control and to the that Kvβ is an to a A in the We and the and the the that of the The is that of NADPH the of a Kvβ indicates that the is and the that Kvβ is a functional AKR. Kvβ Kvβ is a functional AKR to the of the redox reaction Kvβ channel with and channel function and a Kvβ a an and the intracellular of the channel to A and and of and the and the an response not We have the of channel the and the the substrates to the of channel to and and of and the that the substrates channel function through Kvβ. the in channel function is to the redox reaction with the or the the of with the the changes to and and of the of with the changes to and and of a in channel and the of that is in channel when the Kvβ is the studies that the substrates channel functions through and through the redox of the NADPH a Kvβ channel an Kvβ an and when a is to the the NADPH is oxidized. The a in channel hypothesis is can the two and that a with NADPH channel the with the cofactor. the when the with of NADPH of in the and in two of the to the in channel when NADPH to These suggest that Kvβ an can an enzymatic when to its and NADPH to the in channel function as a found that when a and to a channel and a state that not the a in channel can the of channel the as a function of a Since with the NADPH a that channel may also the is channel channel using and in the of the The functions with a of and and channel function to the in the the that channel is the Kvβ Kvβ with the studies the reaction that the of channel can with that of NADPH in a We found that and the NADPH with a of and that in the with the of channel in is that the is a Kvβ is with a Kvβ in an may have properties that in have that Kvβ is a functional and have that Kvβ substrates channel through the NADPH cofactor. These results that the functions of the two dynamically is that of Kvβ with channels channel and properties of channel and the demonstration that Kvβ is a functional is to of channel functions to the of the NADPH as a the the and AKR the is in of the AKR and in Kvβ is an a when the to a has the the a of the to an in an reductase the a of the of the substrates These results that the may an role in and the in Kvβ the of its enzymatic of is and is the the two catalyzes a a of the the of Kvβ is with a the the NADPH a of is Kvβ. These results that Kvβ is a enzyme has a Kvβ a redox that in and channel function or an enzyme is channel here between the two seem to favor the redox The enzymatic the function of Kvβ is to not is to redox the and the with structural studies have that the NADPH a of the the present in the of The is to a the NADPH and the of the in in The of Kvβ that the complex is to to a redox and the channel in a modulated state a of with Kvβ has been in channel and and have found that a the Kvβ-bound NADPH and also channel functions in a not a to the understanding the of the functional coupling and the physiological of functional between the two a of coupling intracellular redox to the excitability of a The results here to understanding the physiological functions of Kvβ. have that Kvβ is a functional and have that Kvβ substrates channel through the NADPH cofactor. These results that the functions of the two dynamically is that of Kvβ with channels channel and properties of channel and the demonstration that Kvβ is a functional is to of channel functions to the of the NADPH as a the the and AKR the is in of the AKR and in Kvβ is an a when the to a has the the a of the to an in an reductase the a of the of the substrates These results that the may an role in and the in Kvβ the of its The enzymatic of is and is the the two catalyzes a a of the the of Kvβ is with a the the NADPH a of is Kvβ. These results that Kvβ is a enzyme has a Kvβ a redox that in and channel function or an enzyme is channel here between the two seem to favor the redox The enzymatic the function of Kvβ is to not is to redox the and the with structural studies have that the NADPH a of the the present in the of The is to a the NADPH and the of the in in The of Kvβ that the complex is to to a redox and the channel in a modulated state a of with Kvβ has been in channel and and have found that a the Kvβ-bound NADPH and also channel functions in a not a to the understanding the of the functional coupling and the physiological of Kvβ. The functional between the two a of coupling intracellular redox to the excitability of a The results here to understanding the physiological functions of Kvβ. We and We AKR and the We and the