Role of different domains in the self-association of rat nucleoporin p62

1994 ◽  
Vol 107 (2) ◽  
pp. 631-638
Author(s):  
F. Buss ◽  
H. Kent ◽  
M. Stewart ◽  
S.M. Bailer ◽  
J.A. Hanover
Keyword(s):  
Electron Microscopy ◽  
Association Studies ◽  
Coiled Coil ◽  
Small Region ◽  
C Terminus ◽  
Linear Chains ◽  
Beta Structure ◽  
Self Association ◽  
And Function

We have expressed rat nucleoporin p62 cDNA in Escherichia coli to obtain material for structural and self-association studies. Electron microscopy and circular dichroism spectroscopy are consistent with a rod-shaped molecule with an alpha-helical coiled-coil domain at its C terminus and a cross-beta structure at its N terminus, separated by a threonine-rich linker, which has a less-defined secondary structure. Electron microscopy and the solubility properties of fragments produced using thrombin and CNBr digestion indicate that p62 molecules associate to form linear chains and that a small region near the C terminus is an important determinant of assembly. This association may have important consequences for pore structure and function; for example, one way p62 could associate would be to form rings in nuclear pores that could function like barrel hoops.

The SPI2-encoded SseA chaperone has discrete domains required for SseB stabilization and export, and binds within the C-terminus of SseB and SseD

Microbiology ◽  
2004 ◽  
Vol 150 (7) ◽  
pp. 2055-2068 ◽  
Author(s):  
Daniel V. Zurawski ◽  
Murry A. Stein
Keyword(s):  
Type Iii Secretion ◽  
Coiled Coil ◽  
Amphipathic Helix ◽  
Yeast Two Hybrid ◽  
C Terminus ◽  
N Terminus ◽  
Domain Mapping ◽  
Self Association ◽  
Discrete Domains ◽  
Two Hybrid

SseA, a key Salmonella virulence determinant, is a small, basic pI protein encoded within the Salmonella pathogenicity island 2 and serves as a type III secretion system chaperone for SseB and SseD. Both SseA partners are subunits of the surface-localized translocon module that delivers effectors into the host cell; SseB is predicted to compose the translocon sheath and SseD is a putative translocon pore subunit. In this study, SseA molecular interactions with its partners were characterized further. Yeast two-hybrid screens indicate that SseA binding requires a C-terminal domain within both partners. An additional central domain within SseD was found to influence binding. The SseA-binding region within SseB was found to encompass a predicted amphipathic helix of a type participating in coiled-coil interactions that are implicated in the assembly of translocon sheaths. Deletions that impinge upon this putative coiled-coiled domain prevent SseA binding, suggesting that SseA occupies a portion of the coiled-coil. SseA occupancy of this motif is envisioned to be sufficient to prevent premature SseB self-association inside bacteria. Domain mapping on the chaperone was also performed. A deletion of the SseA N-terminus, or site-directed mutations within this region, allowed stabilization of SseB, but its export was disrupted. Therefore, the N-terminus of SseA provides a function that is essential for SseB export, but dispensable for partner binding and stabilization.

Role of the C-Terminus of the High-Conductance Calcium-Activated Potassium Channel in Channel Structure and Function

Biochemistry ◽  
2005 ◽  
Vol 44 (30) ◽  
pp. 10135-10144 ◽  
Author(s):  
William A. Schmalhofer ◽  
Manuel Sanchez ◽  
Ge Dai ◽  
Ashvin Dewan ◽  
Lorena Secades ◽  
...  
Keyword(s):  
Potassium Channel ◽  
Structure And Function ◽  
Channel Structure ◽  
C Terminus ◽  
And Function ◽  
Calcium Activated Potassium Channel ◽  
High Conductance

scholarly journals Nbr1 Is a Novel Inhibitor of Ligand-Mediated Receptor Tyrosine Kinase Degradation

2010 ◽  
Vol 30 (24) ◽  
pp. 5672-5685 ◽  
Author(s):  
Faraz K. Mardakheh ◽  
Giulio Auciello ◽  
Tim R. Dafforn ◽  
Joshua Z. Rappoport ◽  
John K. Heath
Keyword(s):  
Tyrosine Kinases ◽  
Protein Localization ◽  
Negative Regulator ◽  
Receptor Internalization ◽  
C Terminus ◽  
Late Endosomes ◽  
Exact Function ◽  
Α Helix ◽  
And Function

ABSTRACT Neighbor of BRCA1 (Nbr1) is a highly conserved multidomain scaffold protein with proposed roles in endocytic trafficking and selective autophagy. However, the exact function of Nbr1 in these contexts has not been studied in detail. Here we investigated the role of Nbr1 in the trafficking of receptor tyrosine kinases (RTKs). We report that ectopic Nbr1 expression inhibits the ligand-mediated lysosomal degradation of RTKs, and this is probably done via the inhibition of receptor internalization. Conversely, the depletion of endogenous NBR1 enhances RTK degradation. Analyses of truncation mutations demonstrated that the C terminus of Nbr1 is essential but not sufficient for this activity. Moreover, the C terminus of Nbr1 is essential but not sufficient for the localization of the protein to late endosomes. We demonstrate that the C terminus of Nbr1 contains a novel membrane-interacting amphipathic α-helix, which is essential for the late endocytic localization of the protein but not for its effect on RTK degradation. Finally, autophagic and late endocytic localizations of Nbr1 are independent of one another, suggesting that the roles of Nbr1 in each context might be distinct. Our results define Nbr1 as a negative regulator of ligand-mediated RTK degradation and reveal the interplay between its various regions for protein localization and function.

Role of the C-terminus and the long cytoplasmic loop in reduced folate carrier expression and function

2002 ◽  
Vol 63 (9) ◽  
pp. 1717-1724 ◽  
Author(s):  
Iraida G. Sharina ◽  
Rongbao Zhao ◽  
Yanhua Wang ◽  
Solomon Babani ◽  
I.David Goldman
Keyword(s):  
Reduced Folate Carrier ◽  
Cytoplasmic Loop ◽  
C Terminus ◽  
And Function

scholarly journals C-terminus of the P4-ATPase ATP8A2 functions in protein folding and regulation of phospholipid flippase activity

2017 ◽  
Vol 28 (3) ◽  
pp. 452-462 ◽  
Author(s):  
Madhavan Chalat ◽  
Kody Moleschi ◽  
Robert S. Molday
Keyword(s):  
Endoplasmic Reticulum ◽  
Atpase Activity ◽  
Deletion Mutants ◽  
Serine Residue ◽  
Cellular Processes ◽  
Phospholipid Asymmetry ◽  
C Terminus ◽  
And Function ◽  
Phospholipid Flippase

ATP8A2 is a P4-ATPase that flips phosphatidylserine and phosphatidylethanolamine across cell membranes. This generates membrane phospholipid asymmetry, a property important in many cellular processes, including vesicle trafficking. ATP8A2 deficiency causes severe neurodegenerative diseases. We investigated the role of the C-terminus of ATP8A2 in its expression, subcellular localization, interaction with its subunit CDC50A, and function as a phosphatidylserine flippase. C-terminal deletion mutants exhibited a reduced tendency to solubilize in mild detergent and exit the endoplasmic reticulum. The solubilized protein, however, assembled with CDC50A and displayed phosphatidylserine flippase activity. Deletion of the C-terminal 33 residues resulted in reduced phosphatidylserine-dependent ATPase activity, phosphatidylserine flippase activity, and neurite extension in PC12 cells. These reduced activities were reversed with 60- and 80-residue C-terminal deletions. Unlike the yeast P4-ATPase Drs2, ATP8A2 is not regulated by phosphoinositides but undergoes phosphorylation on the serine residue within a CaMKII target motif. We propose a model in which the C-terminus of ATP8A2 consists of an autoinhibitor domain upstream of the C-terminal 33 residues and an anti-autoinhibitor domain at the extreme C-terminus. The latter blocks the inhibitory activity of the autoinhibitor domain. We conclude that the C-terminus plays an important role in the efficient folding and regulation of ATP8A2.

Progress in ultrastructural and molecular analysis of the nuclear matrix

1992 ◽  
Vol 50 (1) ◽  
pp. 508-509
Author(s):  
Edward G. Fey
Keyword(s):  
Electron Microscopy ◽  
Nuclear Matrix ◽  
Structure And Function ◽  
Matrix Proteins ◽  
Rna Transcription ◽  
Nuclear Matrix Proteins ◽  
The Past ◽  
And Function

In the past few years, considerable advances have been made regarding the structure and function of the nuclear matrix. In the first half of this presentation, the field of nuclear matrix research will be summarized. Emphasis will be placed on those studies where molecular interactions are demonstrated in situ utilizing high resolution light and/or electron microscopy. Studies demonstrating the role of the nuclear matrix in DNA synthesis and replication, RNA transcription and processing, and the binding of matrix attachment regions to specific nuclear matrix proteins will be summarized.

What microscopic studies of confronting cisternae and annulate lamellae reveal about their similarities and differences

1995 ◽  
Vol 53 ◽  
pp. 926-927
Author(s):  
John R. Palisano ◽  
Karen S. Renzaglia
Keyword(s):  
Electron Microscopy ◽  
Tumor Cells ◽  
Mitotic Cell ◽  
Annulate Lamellae ◽  
Proliferative Stage ◽  
Dividing Cells ◽  
Microscopic Studies ◽  
Similarities And Differences ◽  
And Function

Knowledge of the origin and function of confronting cisternae (CC) and annulate lamellae (AL) has been limited because they are so infrequently observed. CC were first described by Porter in 1955 and have since been observed in a variety of rapidly proliferating tumor cells and selected fetal tissues. AL were first reported in 1952 by McCullough in sea urchin eggs. Although AL have since been observed in a variety of cells, they have been predominately found in oocytes, spermatocytes, and tumor cells. A microscopic study was initiated to gain a better understanding of the origin and role of these two membranous organelles that are frequently found in rapidly dividing cells that lose the ability to express these organelles once their rapid proliferative stage ceases. Because it has proven difficult to interpret the flow of membrane in the mitotic cell by electron microscopy alone, data obtained by electron microscopy was correlated with fluorescence microscopy wherever possible.

Expression in Escherichia coli of fragments of the coiled-coil rod domain of rabbit myosin: influence of different regions of the molecule on aggregation and paracrystal formation

1991 ◽  
Vol 99 (4) ◽  
pp. 823-836
Author(s):  
S.J. Atkinson ◽  
M. Stewart
Keyword(s):  
Escherichia Coli ◽  
Ionic Strength ◽  
Coiled Coil ◽  
Periodic Variation ◽  
Heptad Repeat ◽  
C Terminus ◽  
N Terminus ◽  
Low Ionic Strength ◽  
Myosin Rod

We have expressed in Escherichia coli a cDNA clone corresponding broadly to rabbit light meromyosin (LMM) together with a number of modified polypeptides and have used this material to investigate the role of different aspects of molecular structure on the solubility properties of LMM. The expressed material was characterized biochemically and structurally to ensure that it retained the coiled-coil conformation of the native molecule. Full-length recombinant LMM retained the general solubility properties of myosin and, although soluble at high ionic strength, precipitated when the ionic strength was reduced below 0.3 M. Constructs in which the ‘skip’ residues (that disrupt the coiled-coil heptad repeat) were deleted had solubility properties indistinguishable from the wild type, which indicated that the skip residues did not play a major role in determining the molecular interactions involved in assembly. Deletions from the N terminus of LMM did not alter the solubility properties of the expressed material, but deletion of 92 residues from the C terminus caused a large increase in solubility at low ionic strength, indicating that a determinant important for interaction between LMM molecules was located in this region. The failure of deletions from the molecule's N terminus to alter its solubility radically suggested that the periodic variation of charge along the myosin rod may not be as important as proposed for determining the strength of binding between molecules and thus the solubility of myosin.

scholarly journals TIR-only protein RBA1 recognizes a pathogen effector to regulate cell death inArabidopsis

2017 ◽  
Vol 114 (10) ◽  
pp. E2053-E2062 ◽  
Author(s):  
Marc T. Nishimura ◽  
Ryan G. Anderson ◽  
Karen A. Cherkis ◽  
Terry F. Law ◽  
Qingli L. Liu ◽  
...  
Keyword(s):  
Cell Death ◽  
Immune System ◽  
Pasteurella Multocida ◽  
Coiled Coil ◽  
Nucleotide Binding ◽  
Leucine Rich Repeat ◽  
Multidomain Structure ◽  
Pathogen Effector ◽  
Self Association ◽  
And Function

Detection of pathogens by plants is mediated by intracellular nucleotide-binding site leucine-rich repeat (NLR) receptor proteins. NLR proteins are defined by their stereotypical multidomain structure: an N-terminal Toll–interleukin receptor (TIR) or coiled-coil (CC) domain, a central nucleotide-binding (NB) domain, and a C-terminal leucine-rich repeat (LRR). The plant innate immune system contains a limited NLR repertoire that functions to recognize all potential pathogens. We isolated Response to the bacterial type III effector protein HopBA1 (RBA1), a gene that encodes a TIR-only protein lacking all other canonical NLR domains. RBA1 is sufficient to trigger cell death in response to HopBA1. We generated a crystal structure for HopBA1 and found that it has similarity to a class of proteins that includes esterases, the heme-binding protein ChaN, and an uncharacterized domain ofPasteurella multocidatoxin. Self-association, coimmunoprecipitation with HopBA1, and function of RBA1 require two previously identified TIR–TIR dimerization interfaces. Although previously described as distinct in other TIR proteins, in RBA1 neither of these interfaces is sufficient when the other is disrupted. These data suggest that oligomerization of RBA1 is required for function. Our identification of RBA1 demonstrates that “truncated” NLRs can function as pathogen sensors, expanding our understanding of both receptor architecture and the mechanism of activation in the plant immune system.

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