Bands, interbands and puffs in native Drosophila polytene chromosomes are recognized by a monoclonal antibody to an epitope in the carboxy-terminal tail of histone H1

Chromosoma ◽  
1989 ◽  
Vol 98 (6) ◽  
pp. 411-421 ◽  
Author(s):  
Ronald J. Hill ◽  
Fujiko Watt ◽  
Catherine M. Wilson ◽  
Theodora Fifis ◽  
P. Anne Underwood ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Polytene Chromosomes ◽  
Histone H1 ◽  
Carboxy Terminal

Na+/Ca2+ exchanger in Drosophila: cloning, expression, and transport differences

1997 ◽  
Vol 273 (1) ◽  
pp. C257-C265 ◽  
Author(s):  
A. Ruknudin ◽  
C. Valdivia ◽  
P. Kofuji ◽  
W. J. Lederer ◽  
D. H. Schulze
Keyword(s):  
Protein Kinase ◽  
Protein Function ◽  
Amino Acid Level ◽  
Polytene Chromosomes ◽  
Intracellular Loop ◽  
Loop Region ◽  
Loop Sequence ◽  
Carboxy Terminal ◽  
Gene Maps

cDNAs for the Na+/Ca2+ exchanger from Drosophila melanogaster (Dmel/Nck) have been cloned by homology screening using the human heart Na+/Ca2+ exchanger cDNA. The overall deduced protein structure for Dmel/Nck is similar to that of mammalian Na+/Ca2+ exchanger genes NCX1 and NCX2, having six hydrophobic regions in the amino terminus separated from six at the carboxy-terminal end by a large intracellular loop. Sequence comparison of the Drosophila exchanger cDNAs with NCX1 and NCX2 Na+/Ca2+ exchangers are approximately 46% identical at the deduced amino acid level. Consensus phosphorylation sites for both protein kinase C and protein kinase A are present on the intracellular loop region of the Dmel/Nck. Alternative splicing for the Dmel/Nck gene is suggested in the same intracellular loop region as demonstrated for NCX1. Functionally, the Drosophila Na+/ Ca2+ exchanger expressed in oocytes differs from expressed mammalian NCX1 with regard to Ca2+ transport in Ca2+/ Ca2+ exchange and the effect of monovalent-dependent Ca2+/ Ca2+ exchange. The Dmel/Nck gene maps to chromosome 3 (93A-B) using in situ hybridization to polytene chromosomes, the same position as the Na(+)-K(+)-ATPase, a related transporter. We conclude that, although extracellular Na+ concentration-dependent Ca2+ transport is subserved by both human and Drosophila Na+/Ca2+ exchangers, there are clear and important differences in the transporters, which should be useful in deducing how the Na+/Ca2+ exchanger protein function depends on its structure.

Histone H1 in the centromeric heterochromatin of Glyptotendipes barbipes larval polytene chromosomes

Chromosoma ◽  
1989 ◽  
Vol 98 (1) ◽  
pp. 64-68 ◽  
Author(s):  
S. G. Nonchev ◽  
P. V. Michailova ◽  
C. D. Venkov ◽  
R. G. Tsanev
Keyword(s):  
Polytene Chromosomes ◽  
Histone H1 ◽  
Centromeric Heterochromatin

scholarly journals Tubulin-tyrosine ligase has a binding site on beta-tubulin: a two-domain structure of the enzyme

1987 ◽  
Vol 104 (4) ◽  
pp. 1059-1067 ◽  
Author(s):  
J Wehland ◽  
K Weber
Keyword(s):  
Monoclonal Antibody ◽  
Monoclonal Antibodies ◽  
Enzymatic Activity ◽  
Binding Site ◽  
Gradient Centrifugation ◽  
Beta Tubulin ◽  
Alpha Tubulin ◽  
Alpha Beta ◽  
Tubulin Tyrosine Ligase ◽  
Carboxy Terminal

Tubulin-tyrosine ligase and alpha beta-tubulin form a tight complex which is conveniently monitored by glycerol gradient centrifugation. Using two distinct ligase monoclonal antibodies, several subunit-specific tubulin monoclonal antibodies, and chemical cross-linking, a ligase-binding site was identified on beta-tubulin. This site is retained when the carboxy-terminal domains of both tubulin subunits are removed by subtilisin treatment. The ligase-tubulin complex is also formed when ligase is added to alpha beta-tubulin carrying the monoclonal antibody YL 1/2 which binds only to the carboxyl end of tyrosinated alpha-tubulin. The beta-tubulin-binding site described here explains the extreme substrate specificity of ligase, which does not act on other cellular proteins or carboxy-terminal peptides derived from detyrosinated alpha-tubulin. Differential accessibility of this site in tubulin and in microtubules seems to explain why ligase acts preferentially on unpolymerized tubulin. Ligase exposed to V8-protease is converted to a nicked derivative. This is devoid of enzymatic activity but still forms the complex with tubulin. Gel electrophoresis documents both 30- and a 14-kD domains, each which is immunologically and biochemically distinct and seems to cover the entire molecule. The two domains interact tightly under physiological conditions. The 30-kD domain carries the binding sites for beta-tubulin and ATP. The 14-kD domain can possibly form an additional part of the catalytic site as it harbors the epitope for the monoclonal antibody ID3 which inhibits enzymatic activity but not the formation of the ligase-tubulin complex.

scholarly journals Distribution of ligand-occupied alpha IIb beta 3 in resting and activated human platelets determined by expression of a novel class of ligand-induced binding site recognized by monoclonal antibody AP6

Blood ◽  
1996 ◽  
Vol 88 (3) ◽  
pp. 887-899
Author(s):  
P Nurden ◽  
M Humbert ◽  
RS Piotrowicz ◽  
C Bihour ◽  
C Poujol ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Binding Site ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Gamma Chain ◽  
Rgd Peptides ◽  
Platelet Surface ◽  
Immunogold Staining ◽  
Granule Membrane ◽  
Carboxy Terminal

The sequence beta (3)203–228 is involved, in a yet undetermined manner, in alpha IIb beta 3 function. We now show that murine monoclonal antibody (MoAb) AP6, specific for beta (3)211–221, binds to alpha IIb beta 3 on adenosine diphosphate (ADP)-activated platelets only when the receptor is occupied by intact fibrinogen. The ligand-induced binding- site reported by AP6 is unique in that it is not expressed following occupancy by either RGD peptides or the gamma-chain carboxy-terminal dodecapeptide. Binding of AP6 to platelets coincides temporally with the binding of the MoAb 9F9, specific for a receptor-induced binding site on fibrinogen. Thus, AP6 reports the binding of fibrinogen to the recognition pocket of alpha IIb beta 3. Its binding to thrombin- stimulated washed platelets correlates with secretion as determined using an MoAb to P-selectin. When ultrathin sections of nonactivated platelets were examined by immunogold staining and electron microscopy, AP6 identified a pool of alpha IIb beta 3 colocalizing with P-selectin and suggesting the presence of alpha IIb beta 3-ligand complexes in the alpha-granule membrane. There was little binding of AP6 to surface alpha IIb beta 3 of unstimulated platelets. After ADP-induced activation, AP6 was abundantly distributed over the entire platelet surface, including pseudopods, but only when fibrinogen was present in the medium. ADP had little effect on AP6 reactivity within platelets. This contrasted with washed platelets and thrombin, where extensive AP6 binding was observed within internal membrane pools as early as 10 to 15 seconds after stimulation. Surface labeling with AP6 followed slower kinetics. Flow cytometry on Triton X-100 permeabilized fixed platelets confirmed AP6 binding to alpha IIb beta 3 within the platelet. Thus, our results provide evidence of (1) a pool of alpha-granule alpha IIb beta 3 occupied by ligand in nonactivated platelets; (2) thrombin- induced activation of alpha IIb beta 3 within the platelet, and (3) thrombin-induced mobilization of ligand-bound alpha IIb beta 3 to the surface.

scholarly journals Immunochemical Analysis of UMP Kinase fromEscherichia coli

1999 ◽  
Vol 181 (3) ◽  
pp. 833-840 ◽  
Author(s):  
Stéphanie Landais ◽  
Pierre Gounon ◽  
Christine Laurent-Winter ◽  
Jean-Claude Mazié ◽  
Antoine Danchin ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Polyclonal Antibodies ◽  
Intact Protein ◽  
Enzyme Linked Immunosorbent Assay ◽  
Bacterial Membranes ◽  
E Coli ◽  
Specific Localization ◽  
Ump Kinase ◽  
Carboxy Terminal

ABSTRACT Mono- and polyclonal antibodies directed against UMP kinase fromEscherichia coli were tested with the intact protein or with fragments obtained by deletion mutagenesis. As detected in enzyme-linked immunosorbent assay tests, the carboxy-terminal quarter of UMP kinase is immunodominant. Polyclonal antibodies inhibited the enzyme activity with partial or total loss of allosteric effects exerted by UTP and GTP, respectively. These data indicate that the UTP and GTP binding sites in UMP kinase are only partially overlapping. One monoclonal antibody (44-2) recognized a linear epitope in UMP kinase between residues 171 and 180. A single substitution (D174N) in this segment of the enzyme abolished its interaction with the monoclonal antibody (44-2). Polyclonal antisera were used to identify UMP kinase in the bacterial proteome. The enzyme appears as a single spot on two-dimensional electrophoresis at a pI of 7.24 and an apparent molecular mass of 26 kDa. Immunogold labeling of UMP kinase in wholeE. coli cells shows a localization of the protein near the bacterial membranes. Because the protein does not contain sequences usually required for compartmentalization, the aggregation properties of UMP kinase observed in vitro might play a role in this phenomenon. The specific localization of UMP kinase might also be related to its putative role in cell division.

Calcium Ion-Dependent Monoclonal Antibody Against Human Fibrinogen: Preparation, Characterization, and Application to Fibrinogen Purification

1995 ◽  
Vol 73 (04) ◽  
pp. 662-667 ◽  
Author(s):  
Mikihiro Takebe ◽  
Gilbu Soe ◽  
Isao Kohno ◽  
Teruko Sugo ◽  
Michio Matsuda
Keyword(s):  
Monoclonal Antibody ◽  
Calcium Ion ◽  
Calcium Binding ◽  
Divalent Metal Ions ◽  
Calcium Binding Site ◽  
Human Fibrinogen ◽  
Fibrinogen Binding ◽  
Precipitation Procedure ◽  
Close Proximity ◽  
Carboxy Terminal

SummaryWe have produced a high-affinity monoclonal antibody classified as IgG1 with K-type light chains that recognizes the calcium ion(Ca2+)- dependent conformation of the D-domain of human fibrinogen. Binding of fibrinogen in solution to the insolubilized antibody increased in the presence of increasing concentrations of up to 2 mM Ca2+, the half-maximal binding being reached at 130 μM Ca2+. The dissociation constant was estimated to be 1.6 × 10-8 M at 2 mM Ca2+. The antibody was found also to be dependent on other divalent metal ions including Zn2+, Mn2+, Co2+ and Cu2+, but not Ba2+, Mg2+ or Sr2+. The synthetic Gly-Pro-Arg-Pro-amide peptide, which has recently been shown to bind to close proximity to the calcium binding site in the D-domain, was unable to elicit the conformation for the antigen to be recognized by this antibody. This antibody was found to be a suitable ligand for the immunoaffinity chromatography of normal and abnormal fibrinogens directly from citrated plasma depleted of the vitamin K-dependent proteins or heparinized plasma by eliminating the precipitation procedure widely adopted in conventional techniques of fibrinogen purification. Indeed, fibrinogen Marburg I with the Aa chains depleted of the carboxy-terminal Aα(461-610) residue segment has been purified by this technique, although this dysfibrinogen was difficult to purify by conventional precipitation techniques.

scholarly journals Monoclonal Antibody with Specificity to a Conserved Epitope in the C-Terminal Domain of Histone H1 Variants.

1991 ◽  
Vol 16 (4) ◽  
pp. 323-331 ◽  
Author(s):  
M. Ortiz ◽  
L.A. Bejarano ◽  
M.C. Rendón ◽  
G. Martin ◽  
B.R. Brinkley ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Histone H1 ◽  
Terminal Domain ◽  
Conserved Epitope

scholarly journals A monoclonal antibody that inhibits activation of human Hageman factor (factor XII)

Blood ◽  
1985 ◽  
Vol 65 (1) ◽  
pp. 202-210 ◽  
Author(s):  
EJ Small ◽  
JA Katzmann ◽  
RP Tracy ◽  
OD Ratnoff ◽  
GH Jr Goldsmith ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Binding Activity ◽  
Factor Xii ◽  
Amidolytic Activity ◽  
Hageman Factor ◽  
Amino Terminal ◽  
Terminal Fragment ◽  
Coagulation Assay ◽  
Carboxy Terminal ◽  
Sepharose 4B

Abstract A monoclonal antibody to human Hageman factor (HF, factor XII) was derived from BALB/c mouse spleen cells fused with NS-1 mouse myeloma cells. This antibody, purified from ascites fluid, reacted with HF to inhibit the activation of HF, purified or in normal pooled plasma, as measured by a coagulation assay. The antibody did not inhibit the coagulant activity of activated HF. The antibody also inhibited the generation of amidolytic activity in HF-ellagic acid mixtures, but failed to inhibit the amidolytic properties of the carboxy-terminal fragment of HF (HFf). Amidolytic activity, absent in an HF-monoclonal antibody mixture, was generated upon treatment with insoluble trypsin. Monoclonal antibody, bound to CNBr Sepharose 4B gel (Pharmacia Fine Chemicals, Piscataway, NJ), reversibly bound HF in plasma or in buffer, without activating it. HF was then eluted with 4 mol/L guanidine HCI. The passage of 125I-labeled HF enzymatically cleaved by trypsin through a column of monoclonal antibody-CNBr Sepharose 4B gel resulted in flow- through of HFf with a molecular weight (mol wt) of 30,000 and HF fragments of mol wt 12,000. Elution with 4 mol/L guanidine HCI yielded several HF fragments (mol wt 80,000, 52,000, and 40,000) but not HFf. These data suggest that the single determinant recognized by the murine monoclonal antibody is not on HFf, but rather on the amino-terminal fragment thought to be involved in the binding activity of HF. The monoclonal anti-HF bound to CNBr-activated Sepharose 4B gel could be used to artificially deplete plasma samples of HF.

scholarly journals Phosphorylation of the carboxy-terminal domain of histone H1: effects on secondary structure and DNA condensation

2008 ◽  
Vol 36 (14) ◽  
pp. 4719-4726 ◽  
Author(s):  
Alicia Roque ◽  
Inma Ponte ◽  
José Luis R. Arrondo ◽  
Pedro Suau
Keyword(s):  
Secondary Structure ◽  
Histone H1 ◽  
Dna Condensation ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Carboxy Terminal
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