Conserved Amino Acid Residues within the Amino-terminal Domain of ClpB are Essential for the Chaperone Activity

2002 ◽  
Vol 321 (1) ◽  
pp. 111-120 ◽  
Author(s):  
Zhonghua Liu ◽  
Vekalet Tek ◽  
Vladimir Akoev ◽  
Michal Zolkiewski
Keyword(s):  
Amino Acid ◽  
Chaperone Activity ◽  
Amino Acid Residues ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Amino Terminal Domain

scholarly journals N-Linked Glycosylation at Asn3 and the Positively Charged Residues within the Amino-Terminal Domain of the C1 Inhibitor Are Required for Interaction of the C1 Inhibitor with Salmonella enterica Serovar Typhimurium Lipopolysaccharide and Lipid A

2005 ◽  
Vol 73 (8) ◽  
pp. 4478-4487 ◽  
Author(s):  
Dongxu Liu ◽  
Cort C. Cramer ◽  
Jennifer Scafidi ◽  
Alvin E. Davis
Keyword(s):  
Amino Acid ◽  
Lipid A ◽  
Amino Acid Residues ◽  
C1 Inhibitor ◽  
Gram Negative ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Amino Terminal Domain ◽  
Positively Charged ◽  
Lps Binding

ABSTRACT The C1 inhibitor (C1INH), a plasma complement regulatory protein, prevents endotoxin shock, at least partially via the direct interaction of its amino-terminal heavily glycosylated nonserpin region with gram-negative bacterial lipopolysaccharide (LPS). To further characterize the potential LPS-binding site(s) within the amino-terminal domain, mutations were introduced into C1INH at the three N-linked glycosylation sites and at the four positively charged amino acid residues. A mutant in which Asn3 was replaced with Ala was markedly less effective in its binding to LPS, while substitution of Asn47 or Asn59 had little effect on binding. The mutation of C1INH at all four positively charged amino acid residues (Arg18, Lys22, Lys30, and Lys55) resulted in near-complete failure to interact with LPS. The C1INH mutants that did not bind to LPS also did not suppress LPS binding or LPS-induced up-regulation of tumor necrosis factor alpha mRNA expression in RAW 264.7 macrophages. In addition, the binding of C1INH mutants to diphosphoryl lipid A was decreased in comparison with that of recombinant wild-type C1INH. Therefore, the interaction of C1INH with gram-negative bacterial LPS is dependent both on the N-linked carbohydrate at Asn3 and on the positively charged residues within the amino-terminal domain.

scholarly journals Autoantibodies from patients with primary biliary cirrhosis recognize a restricted region within the cytoplasmic tail of nuclear pore membrane glycoprotein Gp210.

1993 ◽  
Vol 178 (6) ◽  
pp. 2237-2242 ◽  
Author(s):  
R E Nickowitz ◽  
H J Worman
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Primary Biliary Cirrhosis ◽  
Antigenic Determinants ◽  
Nuclear Pore ◽  
Biliary Cirrhosis ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

Patients with primary biliary cirrhosis (PBC) frequently have autoantibodies against a 210-kD integral glycoprotein of the nuclear envelope pore membrane. This protein, termed gp210, has a 1,783-amino acid amino-terminal domain located in the perinuclear space, a 20-amino acid transmembrane segment, and a 58-amino acid cytoplasmic carboxy-terminal tail. We now demonstrate that autoantibodies from 25 patients with PBC that recognize gp210 react with the cytoplasmic carboxy-terminal tail while none react with unmodified linear epitopes in the amino-terminal domain. The epitope(s) recognized by autoantibodies from all 25 patients is contained within a stretch of 15 amino acids. The recognized amino acid sequence is homologous to the protein products of the Escherichia coli mutY gene and Salmonella typhimurium mutB gene with an exact identity of six consecutive amino acids, suggesting that anti-gp210 antibodies may arise by molecular mimicry of bacterial antigenic determinants.

Short related sequences in the cytoplasmic domains of CD4 and CD8 mediate binding to the amino-terminal domain of the p56lck tyrosine protein kinase

1990 ◽  
Vol 10 (5) ◽  
pp. 1853-1862
Author(s):  
A S Shaw ◽  
J Chalupny ◽  
J A Whitney ◽  
C Hammond ◽  
K E Amrein ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Metal Ion ◽  
Cytoplasmic Domain ◽  
Sequence Motif ◽  
Cytoplasmic Domains ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Tyrosine Protein Kinase ◽  
Amino Terminal Domain

We report that the cytoplasmic domains of the T-lymphocyte glycoproteins CD4 and CD8 alpha contain short related amino acid sequences that are involved in binding the amino-terminal domain of the intracellular tyrosine protein kinase, p56lck. Transfer of as few as six amino acid residues from the cytoplasmic domain of the CD8 alpha protein to the cytoplasmic domain of an unrelated protein conferred p56lck binding to the hybrid protein in HeLa cells. The common sequence motif shared by CD4 and CD8 alpha contains two cysteines, and mutation of either cysteine in the CD4 sequence eliminated binding of p56lck.p56lck also contains two cysteine residues within its CD4-CD8 alpha-binding domain, and both are critical to the interaction with CD4 or CD8 alpha. Because the interaction does not involve disulfide bond formation, a metal ion could stabilize the complex.

Assembly properties of altered beta-tubulin polypeptides containing disrupted autoregulatory domains

1989 ◽  
Vol 9 (8) ◽  
pp. 3418-3428
Author(s):  
W Gu ◽  
N J Cowan
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Beta Tubulin ◽  
Alpha Tubulin ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Tubulin Isotype ◽  
The Stability ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

beta-Tubulin synthesis in eucaryotic cells is subject to control by an autoregulatory posttranscriptional mechanism in which the first four amino acids of the beta-tubulin polypeptide act either directly or indirectly to control the stability of beta-tubulin mRNA. To investigate the contribution of this amino-terminal domain to microtubule assembly and dynamics, we introduced a series of deletions encompassing amino acids 2 to 5 of a single mammalian beta-tubulin isotype, M beta 1. Constructs carrying such deletions were inserted into an expression vector, and the ability of the altered polypeptide to coassemble into microtubules was tested by using an anti-M beta 1-specific antibody. We show that the M beta 1 beta-tubulin polypeptide was competent for coassembly into microtubules in transient transfection experiments and in stably transfected cell lines when it lacked either amino acid 2 or amino acids 2 and 3. The capacity of these mutant beta-tubulins to coassemble into polymerized microtubules was only slightly diminished relative to that of unaltered beta-tubulin, and their expression did not influence the viability or growth properties of cell lines carrying these deletions. However, more extensive amino-terminal deletions either severely compromised or abolished the capacity for coassembly. In analogous experiments in which alterations were introduced into the amino-terminal domain of a mammalian alpha-tubulin isotype, M alpha 4, deletion of amino acid 2 did not affect the ability of the altered polypeptide to coassemble, although removal of additional amino-terminal residues essentially abolished the capacity for competent coassembly. The stability of the altered assembly-competent alpha- and beta-tubulin polypeptides was measured in pulse-chase experiments and found to be indistinguishable from the stability of the corresponding unaltered polypeptides. An assembly-competent M alpha 4 polypeptide carrying a deletion encompassing the 12 carboxy-terminal amino acids also had a half-life indistinguishable from that of the wild-type alpha-tubulin molecule. These data suggest that the universally conserved amino terminus of beta-tubulin acts largely in a regulatory role and that the carboxy-terminal domain of alpha-tubulin is not essential for coassembly in mammalian cells in vivo.

scholarly journals Assembly properties of altered beta-tubulin polypeptides containing disrupted autoregulatory domains.

1989 ◽  
Vol 9 (8) ◽  
pp. 3418-3428 ◽  
Author(s):  
W Gu ◽  
N J Cowan
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Beta Tubulin ◽  
Alpha Tubulin ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Tubulin Isotype ◽  
The Stability ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

beta-Tubulin synthesis in eucaryotic cells is subject to control by an autoregulatory posttranscriptional mechanism in which the first four amino acids of the beta-tubulin polypeptide act either directly or indirectly to control the stability of beta-tubulin mRNA. To investigate the contribution of this amino-terminal domain to microtubule assembly and dynamics, we introduced a series of deletions encompassing amino acids 2 to 5 of a single mammalian beta-tubulin isotype, M beta 1. Constructs carrying such deletions were inserted into an expression vector, and the ability of the altered polypeptide to coassemble into microtubules was tested by using an anti-M beta 1-specific antibody. We show that the M beta 1 beta-tubulin polypeptide was competent for coassembly into microtubules in transient transfection experiments and in stably transfected cell lines when it lacked either amino acid 2 or amino acids 2 and 3. The capacity of these mutant beta-tubulins to coassemble into polymerized microtubules was only slightly diminished relative to that of unaltered beta-tubulin, and their expression did not influence the viability or growth properties of cell lines carrying these deletions. However, more extensive amino-terminal deletions either severely compromised or abolished the capacity for coassembly. In analogous experiments in which alterations were introduced into the amino-terminal domain of a mammalian alpha-tubulin isotype, M alpha 4, deletion of amino acid 2 did not affect the ability of the altered polypeptide to coassemble, although removal of additional amino-terminal residues essentially abolished the capacity for competent coassembly. The stability of the altered assembly-competent alpha- and beta-tubulin polypeptides was measured in pulse-chase experiments and found to be indistinguishable from the stability of the corresponding unaltered polypeptides. An assembly-competent M alpha 4 polypeptide carrying a deletion encompassing the 12 carboxy-terminal amino acids also had a half-life indistinguishable from that of the wild-type alpha-tubulin molecule. These data suggest that the universally conserved amino terminus of beta-tubulin acts largely in a regulatory role and that the carboxy-terminal domain of alpha-tubulin is not essential for coassembly in mammalian cells in vivo.

scholarly journals THE STRUCTURAL BASIS FOR BINDING OF COMPLEMENT BY IMMUNOGLOBULIN M

1974 ◽  
Vol 140 (4) ◽  
pp. 1117-1121 ◽  
Author(s):  
Mary M. Hurst ◽  
John E. Volanakis ◽  
Raymond B. Hester ◽  
Robert M. Stroud ◽  
J. Claude Bennett
Keyword(s):  
Amino Acid ◽  
Structural Features ◽  
Immunoglobulin M ◽  
Structural Basis ◽  
Amino Acid Residues ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Carboxy Terminal ◽  
Insight Into

An insight into the structural features of human IgM that are responsible for its capacity to bind the first component of complement (C) has been obtained by examining the ability of IgM subfragments to bind active C1 (C1). The smallest two fragments found to bind C1 were the major CNBr fragment of the Fc portion of IgM and the CH4 fragment of the carboxy-terminal domain. The smallest fragment which fixes C1 has a disaggregated mol wt of 6,800, consists of 60 residues, and contains no carbohydrate. Structural considerations and sequence overlaps suggest that the amino-terminal side of the CH4 domain (24 amino acid residues) might be responsible for fixing C1.

scholarly journals Short related sequences in the cytoplasmic domains of CD4 and CD8 mediate binding to the amino-terminal domain of the p56lck tyrosine protein kinase.

1990 ◽  
Vol 10 (5) ◽  
pp. 1853-1862 ◽  
Author(s):  
A S Shaw ◽  
J Chalupny ◽  
J A Whitney ◽  
C Hammond ◽  
K E Amrein ◽  
...  
Keyword(s):  
Amino Acid ◽  
Protein Kinase ◽  
Metal Ion ◽  
Cytoplasmic Domain ◽  
Sequence Motif ◽  
Cytoplasmic Domains ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Tyrosine Protein Kinase ◽  
Amino Terminal Domain

We report that the cytoplasmic domains of the T-lymphocyte glycoproteins CD4 and CD8 alpha contain short related amino acid sequences that are involved in binding the amino-terminal domain of the intracellular tyrosine protein kinase, p56lck. Transfer of as few as six amino acid residues from the cytoplasmic domain of the CD8 alpha protein to the cytoplasmic domain of an unrelated protein conferred p56lck binding to the hybrid protein in HeLa cells. The common sequence motif shared by CD4 and CD8 alpha contains two cysteines, and mutation of either cysteine in the CD4 sequence eliminated binding of p56lck.p56lck also contains two cysteine residues within its CD4-CD8 alpha-binding domain, and both are critical to the interaction with CD4 or CD8 alpha. Because the interaction does not involve disulfide bond formation, a metal ion could stabilize the complex.

scholarly journals Two distinct domains in the yeast transcription factor IID and evidence for a TATA box-induced conformational change.

1991 ◽  
Vol 11 (1) ◽  
pp. 63-74 ◽  
Author(s):  
P M Lieberman ◽  
M C Schmidt ◽  
C C Kao ◽  
A J Berk
Keyword(s):  
Transcription Factor ◽  
Amino Acid ◽  
Dna Binding ◽  
Conformational Change ◽  
Tata Box ◽  
Amino Terminus ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

Transcription factor IID from Saccharomyces cerevisiae (YIID) binds the TATA box element present in most RNA polymerase II promoters. In this work, partial proteolysis was used as a biochemical probe of YIID structure. YIID consists of a protease-sensitive amino terminus and a highly stable, protease-resistant carboxy-terminal core. The cleavage sites of the predominant chymotrypsin- and trypsin-derived fragments were mapped to amino acid residues 40 to 41 and 48 to 49, respectively, by amino-terminal peptide sequencing. Removal of the amino terminus resulted in a dramatic increase in the ability of YIID to form a stable complex with DNA during gel electrophoresis mobility shift assays and a two- to fourfold increase in DNA-binding affinity, as assayed by DNase I footprinting analysis. The carboxy-terminal 190-amino-acid core was competent for transcription in vitro and was similar in activity to native YIID. DNA containing a TATA element induced hypersensitive sites in the amino-terminal domain and stabilized the core domain to further proteolytic attack. Native YIID did not bind to a TATA box at 0 degrees C, whereas the carboxy-terminal DNA-binding domain did. These results suggest that YIID undergoes a conformational change upon binding to a TATA box. Southern blotting showed that the carboxy-terminal domain is highly conserved, while the amino-terminal domain diverged rapidly in evolution, even between closely related budding yeasts.

Two distinct domains in the yeast transcription factor IID and evidence for a TATA box-induced conformational change

1991 ◽  
Vol 11 (1) ◽  
pp. 63-74
Author(s):  
P M Lieberman ◽  
M C Schmidt ◽  
C C Kao ◽  
A J Berk
Keyword(s):  
Transcription Factor ◽  
Amino Acid ◽  
Dna Binding ◽  
Conformational Change ◽  
Tata Box ◽  
Amino Terminus ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Carboxy Terminal ◽  
Amino Terminal Domain

Transcription factor IID from Saccharomyces cerevisiae (YIID) binds the TATA box element present in most RNA polymerase II promoters. In this work, partial proteolysis was used as a biochemical probe of YIID structure. YIID consists of a protease-sensitive amino terminus and a highly stable, protease-resistant carboxy-terminal core. The cleavage sites of the predominant chymotrypsin- and trypsin-derived fragments were mapped to amino acid residues 40 to 41 and 48 to 49, respectively, by amino-terminal peptide sequencing. Removal of the amino terminus resulted in a dramatic increase in the ability of YIID to form a stable complex with DNA during gel electrophoresis mobility shift assays and a two- to fourfold increase in DNA-binding affinity, as assayed by DNase I footprinting analysis. The carboxy-terminal 190-amino-acid core was competent for transcription in vitro and was similar in activity to native YIID. DNA containing a TATA element induced hypersensitive sites in the amino-terminal domain and stabilized the core domain to further proteolytic attack. Native YIID did not bind to a TATA box at 0 degrees C, whereas the carboxy-terminal DNA-binding domain did. These results suggest that YIID undergoes a conformational change upon binding to a TATA box. Southern blotting showed that the carboxy-terminal domain is highly conserved, while the amino-terminal domain diverged rapidly in evolution, even between closely related budding yeasts.

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