Molecular Cloning and Expression of the Novel Splice Variants of K+ Channel-Interacting Protein 2

2001 ◽  
Vol 282 (1) ◽  
pp. 96-102 ◽  
Author(s):  
Susumu Ohya ◽  
Yuichi Morohashi ◽  
Katsuhiko Muraki ◽  
Taisuke Tomita ◽  
Minoru Watanabe ◽  
...  
Keyword(s):  
Molecular Cloning ◽  
Splice Variants ◽  
Cloning And Expression ◽  
K Channel ◽  
The Novel ◽  
Interacting Protein

Functional properties of a brain-specific NH2-terminally spliced modulator of Kv4 channels

2003 ◽  
Vol 285 (1) ◽  
pp. C161-C170 ◽  
Author(s):  
Linda M. Boland ◽  
Min Jiang ◽  
So Yeong Lee ◽  
Scott C. Fahrenkrug ◽  
Mark T. Harnett ◽  
...  
Keyword(s):  
Terminal Region ◽  
K Channel ◽  
The Novel ◽  
Splice Form ◽  
Interacting Protein ◽  
Current Decay ◽  
Amino Terminal ◽  
Human Genes ◽  
Kv4 Channels ◽  
Inactivation Gating

Kv4/K channel-interacting protein (KChIP) potassium channels are a major class of rapidly inactivating K channels in brain and heart. Considering the importance of alternative splicing to the quantitative features of KChIP gating modulation, a previously uncharacterized splice form of KChIP1 was functionally characterized. The KChIP1b splice variant differs from the previously characterized KChIP1a splice form by the inclusion of a novel amino-terminal region that is encoded by an alternative exon that is conserved in mouse, rat, and human genes. The expression of KChIP1b mRNA was high in brain but undetectable in heart or liver by RT-PCR. In cerebellar tissue, KChIP1b and KChIP1a transcripts were expressed at nearly equal levels. Coexpression of KChIP1b or KChIP1a with Kv4.2 channels in oocytes slowed K current decay and destabilized open-inactivated channel gating. Like other KChIP subunits, KChIP1b increased Kv4.2 current amplitude and KChIP1b also shifted Kv4.2 conductance-voltage curves by —10 mV. The development of Kv4.2 channel inactivation accessed from closed gating states was faster with KChIP1b coexpression. Deletion of the novel amino-terminal region in KChIP1b selectively altered the subunit's modulation of Kv4.2 closed inactivation gating. The role of the KChIP1b NH2-terminal region was further confirmed by direct comparison of the properties of the NH2-terminal deletion mutant and the KChIP1a subunit, which is encoded by a transcript that lacks the novel exon. The features of KChIP1b modulation of Kv4 channels are likely to be conserved in mammals and demonstrate a role for the KChIP1 NH2-terminal region in the regulation of closed inactivation gating.

scholarly journals Molecular cloning and expression of a fifth muscarinic acetylcholine receptor

1989 ◽  
Vol 264 (13) ◽  
pp. 7328-7337
Author(s):  
C F Liao ◽  
A P Themmen ◽  
R Joho ◽  
C Barberis ◽  
M Birnbaumer ◽  
...  
Keyword(s):  
Molecular Cloning ◽  
Acetylcholine Receptor ◽  
Muscarinic Acetylcholine Receptor ◽  
Cloning And Expression ◽  
Muscarinic Acetylcholine

scholarly journals Split NanoLuc technology allows quantitation of interactions between PII protein and its receptors with unprecedented sensitivity and reveals transient interactions

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rokhsareh Rozbeh ◽  
Karl Forchhammer
Keyword(s):  
The Novel ◽  
Multiple Target ◽  
Interacting Protein ◽  
Effector Molecule ◽  
Target Proteins ◽  
Pii Protein ◽  
Protein X ◽  
Cellular Context ◽  
Pii Proteins ◽  
Transient Interactions

AbstractPII proteins constitute a widespread signal transduction superfamily in the prokaryotic world. The canonical PII signal proteins sense metabolic state of the cells by binding the metabolite molecules ATP, ADP and 2-oxoglutarate. Depending on bound effector molecule, PII proteins interact with and modulate the activity of multiple target proteins. To investigate the complexity of interactions of PII with target proteins, analytical methods that do not disrupt the native cellular context are required. To this purpose, split luciferase proteins have been used to develop a novel complementation reporter called NanoLuc Binary Technology (NanoBiT). The luciferase NanoLuc is divided in two subunits: a 18 kDa polypeptide termed “Large BiT” and a 1.3 kDa peptide termed “Small BiT”, which only weakly associate. When fused to proteins of interest, they reconstitute an active luciferase when the proteins of interest interact. Therefore, we set out to develop a new NanoBiT sensor based on the interaction of PII protein from Synechocystis sp. PCC6803 with PII-interacting protein X (PipX) and N-acetyl-L-glutamate kinase (NAGK). The novel NanoBiT sensor showed unprecedented sensitivity, which made it possible to detect even weak and transient interactions between PII variants and their interacting partners, thereby shedding new light in PII signalling processes.

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