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Daniel L Minor

Publications and source records attributed to Daniel L Minor.

11 recordsLinked to original sources
PubMed2006

Three-dimensional structure of the KChIP1-Kv4.3 T1 complex reveals a cross-shaped octamer.

Brain I(A) and cardiac I(to) currents arise from complexes containing Kv4 voltage-gated potassium channels and cytoplasmic calcium-sensor proteins (KChIPs). Here, we present X-ray crystallographic and small-angle X-ray scattering data that show that the KChIP1-Kv4.3 N-terminal cytoplasmic domain complex is a cross-shaped octamer bearing two principal interaction sites. Site 1 comprises interactions between a unique Kv4 channel N-terminal hydrophobic segment and a hydrophobic pocket formed by displacement of the KChIP H10 helix. Site 2 comprises interactions between a T1 assembly domain loop and the KChIP H2 helix. Functional and biochemical studies indicate that site 1 influences channel trafficking, whereas site 2 affects channel gating, and that calcium binding is intimately linked to KChIP folding and complex formation. Together, the data resolve how Kv4 channels and KChIPs interact and provide a framework for understanding how KChIPs modulate Kv4 function.

Amino Acid Sequence↗
PubMed2006

Coiled coils direct assembly of a cold-activated TRP channel.

Transient receptor potential (TRP) channels mediate numerous sensory transduction processes and are thought to function as tetramers. TRP channel physiology is well studied; however, comparatively little is understood regarding TRP channel assembly. Here, we identify an autonomously folded assembly domain from the cold- and menthol-gated channel TRPM8. We show that the TRPM8 cytoplasmic C-terminal domain contains a coiled coil that is necessary for channel assembly and sufficient for tetramer formation. Cell biological experiments indicate that coiled-coil formation is required for proper channel maturation and trafficking and that the coiled-coil domain alone can act as a dominant-negative inhibitor of functional channel expression. Our data define an authentic TRP modular assembly domain, establish a clear role for coiled coils in ion channel assembly, demonstrate that coiled-coil assembly domains are a general feature of TRPM channels, and delineate a new tool that should be of general use in dissecting TRPM channel function.

Amino Acid Sequence↗
PubMed2006

A multimeric membrane protein reveals 14-3-3 isoform specificity in forward transport in yeast.

Arginine (Arg)-based endoplasmic reticulum (ER) localization signals are sorting motifs involved in the quality control of multimeric membrane proteins. They are distinct from other ER localization signals like the C-terminal di-lysine [-K(X)KXX] signal. The Pmp2p isoproteolipid, a type I yeast membrane protein, reports faithfully on the activity of sorting signals when fused to a tail containing either an Arg-based motif or a -KKXX signal. This reporter reveals that the Arg-based ER localization signals from mammalian Kir6.2 and GB1 proteins are functional in yeast. Thus, the machinery involved in recognition of Arg-based signals is evolutionarily conserved. Multimeric presentation of the Arg-based signal from Kir6.2 on Pmp2p results in forward transport, which requires 14-3-3 proteins encoded in yeast by BMH1 and BMH2 in two isoforms. Comparison of a strain without any 14-3-3 proteins (Deltabmh2) and the individual Deltabmh1 or Deltabmh2 shows that the role of 14-3-3 in the trafficking of this multimeric Pmp2p reporter is isoform-specific. Efficient forward transport requires the presence of Bmh1p. The specific role of Bmh1p is not due to differences in abundance or affinity between the isoforms. Our results imply that 14-3-3 proteins mediate forward transport by a mechanism distinct from simple masking of the Arg-based signal.

14-3-3 Proteins↗
PubMed2005

Insights into voltage-gated calcium channel regulation from the structure of the CaV1.2 IQ domain-Ca2+/calmodulin complex.

Changes in activity-dependent calcium flux through voltage-gated calcium channels (Ca(V)s) drive two self-regulatory calcium-dependent feedback processes that require interaction between Ca(2+)/calmodulin (Ca(2+)/CaM) and a Ca(V) channel consensus isoleucine-glutamine (IQ) motif: calcium-dependent inactivation (CDI) and calcium-dependent facilitation (CDF). Here, we report the high-resolution structure of the Ca(2+)/CaM-Ca(V)1.2 IQ domain complex. The IQ domain engages hydrophobic pockets in the N-terminal and C-terminal Ca(2+)/CaM lobes through sets of conserved 'aromatic anchors.' Ca(2+)/N lobe adopts two conformations that suggest inherent conformational plasticity at the Ca(2+)/N lobe-IQ domain interface. Titration calorimetry experiments reveal competition between the lobes for IQ domain sites. Electrophysiological examination of Ca(2+)/N lobe aromatic anchors uncovers their role in Ca(V)1.2 CDF. Together, our data suggest that Ca(V) subtype differences in CDI and CDF are tuned by changes in IQ domain anchoring positions and establish a framework for understanding CaM lobe-specific regulation of Ca(V)s.

Amino Acid Motifs↗
PubMed2005

The pore helix dipole has a minor role in inward rectifier channel function.

Ion channels lower the energetic barrier for ion passage across cell membranes and enable the generation of bioelectricity. Electrostatic interactions between permeant ions and channel pore helix dipoles have been proposed as a general mechanism for facilitating ion passage. Here, using genetic selections to probe interactions of an exemplar potassium channel blocker, barium, with the inward rectifier Kir2.1, we identify mutants bearing positively charged residues in the potassium channel signature sequence at the pore helix C terminus. We show that these channels are functional, selective, resistant to barium block, and have minimally altered conductance properties. Both the experimental data and model calculations indicate that barium resistance originates from electrostatics. We demonstrate that potassium channel function is remarkably unperturbed when positive charges occur near the permeant ions at a location that should counteract pore helix electrostatic effects. Thus, contrary to accepted models, the pore helix dipole seems to be a minor factor in potassium channel permeation.

Animals↗
PubMed2005

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Protein Structure, Secondary↗
PubMed2004

Structure of a complex between a voltage-gated calcium channel beta-subunit and an alpha-subunit domain.

Voltage-gated calcium channels (Ca(V)s) govern muscle contraction, hormone and neurotransmitter release, neuronal migration, activation of calcium-dependent signalling cascades, and synaptic input integration. An essential Ca(V) intracellular protein, the beta-subunit (Ca(V)beta), binds a conserved domain (the alpha-interaction domain, AID) between transmembrane domains I and II of the pore-forming alpha(1) subunit and profoundly affects multiple channel properties such as voltage-dependent activation, inactivation rates, G-protein modulation, drug sensitivity and cell surface expression. Here, we report the high-resolution crystal structures of the Ca(V)beta2a conserved core, alone and in complex with the AID. Previous work suggested that a conserved region, the beta-interaction domain (BID), formed the AID-binding site; however, this region is largely buried in the Ca(V)beta core and is unavailable for protein-protein interactions. The structure of the AID-Ca(V)beta2a complex shows instead that Ca(V)beta2a engages the AID through an extensive, conserved hydrophobic cleft (named the alpha-binding pocket, ABP). The ABP-AID interaction positions one end of the Ca(V)beta near the intracellular end of a pore-lining segment, called IS6, that has a critical role in Ca(V) inactivation. Together, these data suggest that Ca(V)betas influence Ca(V) gating by direct modulation of IS6 movement within the channel pore.

Amino Acid Sequence↗
PubMed2003

The structure of the APPBP1-UBA3-NEDD8-ATP complex reveals the basis for selective ubiquitin-like protein activation by an E1.

E1 enzymes initiate ubiquitin-like protein (ubl) transfer cascades by catalyzing adenylation of the ubl's C terminus. An E1's selectivity for its cognate ubl is essential because the E1 subsequently coordinates the ubl with its correct downstream pathway. We report here the structure of the 120 kDa quaternary complex between human APPBP1-UBA3, a heterodimeric E1, its ubl NEDD8, and ATP. The E1 selectively recruits NEDD8 through a bipartite interface, involving a domain common to all ubl activating enzymes including bacterial ancestors, and also eukaryotic E1-specific sequences. By modeling ubiquitin into the NEDD8 binding site and performing mutational analysis, we identify a single conserved arginine in APPBP1-UBA3 that acts as a selectivity gate, preventing misactivation of ubiquitin by NEDD8's E1. NEDD8 residues that interact with E1 correspond to residues in ubiquitin important for binding the proteasome and other ubiquitin-interacting proteins, suggesting that the conjugation and recognition machineries have coevolved for each specific ubl.

Adenosine Triphosphate↗
PubMed2002

Consensus-derived structural determinants of the ankyrin repeat motif.

The ankyrin repeat is one of the most common, modular, protein-protein interaction motifs in nature. To understand the structural determinants of this family of proteins and extract the consensus information that defines the architecture of this motif, we have designed a series of idealized ankyrin repeat proteins containing one, two, three, or four repeats by using statistical analysis of approximately 4,000 ankyrin repeat sequences from the PFAM database. Biophysical and x-ray crystallographic studies of the three and four repeat constructs (3ANK and 4ANK) to 1.26 and 1.5 A resolution, respectively, demonstrate that these proteins are well-folded, monomeric, display high thermostability, and adopt a very regular, tightly packed ankyrin repeat fold. Mapping the degree of amino acid conservation at each position on the 4ANK structure shows that most nonconserved residues are clustered on the surface of the molecule that has been designated as the binding site in naturally occurring ankyrin repeat proteins. Thus, the consensus amino acid sequence contains all information required to define the ankyrin repeat fold. Our results suggest that statistical analysis and the consensus sequence approach can be used as an effective method to design proteins with complex topologies. These generic ankyrin repeat proteins can serve as prototypes for dissecting the rules of molecular recognition mediated by ankyrin repeats and for engineering proteins with novel biological functions.

Amino Acid Sequence↗