scholarly journals Mechanical Regulation of a Cell Binding Site: Bacterial Adhesins can Distinguish Different Physical States of Fibronectin

2010 ◽  
Vol 98 (3) ◽  
pp. 242a
Author(s):  
Samuel Hertig
Keyword(s):  
Binding Site ◽  
Cell Binding ◽  
Bacterial Adhesins ◽  
A Cell ◽  
Mechanical Regulation

Structural features and functional domains of amassin-1, a cell-binding olfactomedin protein

2007 ◽  
Vol 85 (5) ◽  
pp. 552-562 ◽  
Author(s):  
Brian J. Hillier ◽  
Victor D. Vacquier
Keyword(s):  
Disulfide Bonds ◽  
Biological Activities ◽  
Structural Features ◽  
Body Cavity ◽  
Binding Activity ◽  
Coiled Coils ◽  
Cell Binding ◽  
Calcium Dependent ◽  
Dependent Cell ◽  
A Cell

Amassin-1 mediates a rapid cell adhesion that tightly adheres sea urchin coelomocytes (body cavity immunocytes) together. Three major structural regions exist in amassin-1: a short β region, 3 coiled coils, and an olfactomedin domain. Amassin-1 contains 8 disulfide-bonded cysteines that, upon reduction, render it inactive. Truncated forms of recombinant amassin-1 were expressed and purified from Pichia pastoris and their disulfide bonding and biological activities investigated. Expressed alone, the olfactomedin domain contained 2 intramolecular disulfide bonds, existed in a monomeric state, and inhibited amassin-1-mediated clotting of coelomocytes by a calcium-dependent cell-binding activity. The N-terminal β region, containing 3 cysteines, was not required for clotting activity. The coiled coils may dimerize amassin-1 in a parallel orientation through a homodimerizing disulfide bond. Neither amassin-1 fragments that were disulfide-linked as dimers or that were engineered to exist as dimers induced coelomocytes clotting. Clotting required higher multimeric states of amassin-1, possibly tetramers, which occurred through the N-terminal β region and (or) the first segment of coiled coils.

Vitamin K epoxide reductase significantly improves carboxylation in a cell line overexpressing factor X

Blood ◽  
2005 ◽  
Vol 106 (12) ◽  
pp. 3811-3815 ◽  
Author(s):  
Yan-Mei Sun ◽  
Da-Yun Jin ◽  
Rodney M. Camire ◽  
Darrel W. Stafford
Keyword(s):  
Binding Site ◽  
Cell Line ◽  
Vitamin K ◽  
Turnover Rate ◽  
Factor X ◽  
Vitamin K Epoxide Reductase ◽  
Transfected Cells ◽  
Epoxide Reductase ◽  
A Cell ◽  
Practical Implications

Previously we reported that we could increase the fraction of carboxylated factor X by reducing the affinity of the propeptide for its binding site on human gamma glutamyl carboxylase. We attributed this to an increased turnover rate. However, even with the reduced affinity propeptide, when sufficient overproduction of factor X is achieved, there is still a significant fraction of uncarboxylated recombinant factor X. We report here that the factor X of such a cell line was only 52% carboxylated but that the fraction of carboxylated factor X could be increased to 92% by coexpressing the recently identified gene for vitamin K epoxide reductase. Because vitamin K is in excess in both the untransfected and vitamin K epoxide reductase (VKOR)–transfected cells, the simplest explanation for this result is that VKOR catalyzes both the reduction of vitamin K epoxide to vitamin K and the conversion of vitamin K to vitamin K hydroquinone. In addition to its mechanistic relevance, this observation has practical implications for overproducing recombinant vitamin K–dependent proteins for therapeutic use.

scholarly journals Mapping of a cell-binding domain in the cell adhesion molecule gp80 of Dictyostelium discoideum.

1988 ◽  
Vol 107 (5) ◽  
pp. 1835-1843 ◽  
Author(s):  
R K Kamboj ◽  
L M Wong ◽  
T Y Lam ◽  
C H Siu
Keyword(s):  
Cell Adhesion ◽  
Dictyostelium Discoideum ◽  
Fusion Proteins ◽  
Binding Activity ◽  
Binding Domain ◽  
Globular Domain ◽  
Cell Binding ◽  
Homophilic Interaction ◽  
A Cell ◽  
Cell Binding Domain

At the aggregation stage of Dictyostelium discoideum development, a cell surface glycoprotein of Mr 80,000 (gp80) has been found to mediate the EDTA-resistant type of cell-cell adhesion via homophilic interaction (Siu, C.-H., A. Cho, and A. H. C. Choi. 1987. J. Cell Biol. 105:2523-2533). To investigate the structure-function relationships of gp80, we have isolated full length cDNA clones for gp80 and determined the DNA sequence. The deduced structure of gp80 showed three major domains. An amino-terminal globular domain composed of the bulk of the protein is supported by a short stalk region, which is followed by a membrane anchor at the carboxy terminus. Structural analysis suggested that the cell-binding domain of gp80 resides within the globular domain near the amino terminus. To investigate the relationship of the cell-binding activity to this region of the polypeptide, three protein A/gp80 (PA80) gene fusions were constructed using the expression vector pRIT2T. These PA80 fusion proteins were assayed for their ability to bind to aggregation stage cells. Binding of 125I-labeled fusion proteins PA80I (containing the Val123 to Ile514 fragment of gp80) and PA80II (Val123 to Ala258) was dosage dependent and could be inhibited by precoating cells with the cell cohesion-blocking mAb 80L5C4. On the other hand, there was no appreciable binding of PA80III (Ile174 to Ile514) to cells. Reassociation of cells was significantly inhibited in the presence of PA80I or PA80II. In addition, 125I-labeled PA80II exhibited homophilic interaction with immobilized PA80I, PA80II, or gp80. The results of these studies lead to the mapping of a cell-binding domain in the region between Val123 and Leu173 of gp80 and provide direct evidence that the cell-binding activity of gp80 resides in the protein moiety.

In Vitro Removal of Ochratoxin A by Wine Lactic Acid Bacteria

2007 ◽  
Vol 70 (9) ◽  
pp. 2155-2160 ◽  
Author(s):  
VINCENZO DEL PRETE ◽  
HECTOR RODRIGUEZ ◽  
ALFONSO V. CARRASCOSA ◽  
BLANCA de las RIVAS ◽  
EMILIA GARCIA-MORUNO ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Ochratoxin A ◽  
High Performance ◽  
Culture Medium ◽  
Degradation Products ◽  
Cell Binding ◽  
A Cell ◽  
In Vitro Interaction

A study was carried out to determine the in vitro interaction between ochratoxin A (OTA) and wine lactic acid bacteria (LAB). Fifteen strains belonging to five relevant oenological LAB species were grown in liquid synthetic culture medium containing OTA. The portion of OTA removed during the bacterial growth was 8 to 28%. The OTA removed from the supernatants was partially recovered (31 to 57%) from the bacterial pellet. Cell-free extracts of three representative strains were produced by disrupting cells in a French pressure cell. The ability of crude cell-free extracts to degrade OTA was studied. OTA was not degraded by cell-free extracts of wine LAB strains, and no degradation products of OTA were detected in the high-performance liquid chromatograms of the methanol extract of the bacterial pellet. On the basis of these results, we conclude that OTA removal by wine LAB is a cell-binding phenomenon. The chemistry and the molecular basis of OTA binding to wine LAB remains unknown.

In vitro incorporation of a cell-binding protein to a lentiviral vector using an engineered split intein enables targeted delivery of genetic cargo

2015 ◽  
Vol 112 (12) ◽  
pp. 2611-2617 ◽  
Author(s):  
Ana M. Chamoun-Emanuelli ◽  
Gus Wright ◽  
Smith Roger ◽  
Robert C. Münch ◽  
Christian J. Buchholz ◽  
...  
Keyword(s):  
Targeted Delivery ◽  
Lentiviral Vector ◽  
Binding Protein ◽  
Cell Binding ◽  
Split Intein ◽  
A Cell

scholarly journals Bioinspired Design of Lysolytic Triterpenoid-peptide Conjugates that Kill African Trypanosomes

2018 ◽  
Author(s):  
W.-Matthias Leeder ◽  
Fabian Giehler ◽  
Juliane Joswig ◽  
H. Ulrich Göringer
Keyword(s):  
Protein Complexes ◽  
Building Blocks ◽  
Optimized Design ◽  
Natural Immunity ◽  
Cell Binding ◽  
Peptide Conjugates ◽  
African Trypanosomes ◽  
Stepwise Development ◽  
A Cell ◽  
The Many

AbstractHumans have evolved a natural immunity against Trypanosoma brucei infections, which is executed by two serum (lipo)protein complexes known as trypanolytic factors (TLF). Active TLF-ingredient is the primate-specific apolipoprotein L1 (ApoL1). The protein has a pore-forming activity that kills parasites by lysosomal and mitochondrial membrane fenestration. Of the many trypanosome subspecies only two are able to counteract the activity of ApoL1, which illustrates its evolutionary optimized design and trypanocidal potency. Here we ask the question whether a synthetic (syn)TLF can be synthesized using the design principles of the natural TLF-complexes but relying on different chemical building blocks. We demonstrate the stepwise development of triterpenoid-peptide conjugates, in which the triterpenoids act as a cell binding, uptake and lysosomal transport-moduls and the synthetic peptide GALA as a pH-sensitive, pore-forming lysolytic toxin. As designed, the conjugate kills infective-stage African trypanosomes through lysosomal lysis demonstrating proof-of-principle for the bioinspired, forward-design of a synTLF.

scholarly journals Characterization of a type IV collagen major cell binding site with affinity to the alpha 1 beta 1 and the alpha 2 beta 1 integrins.

1991 ◽  
Vol 113 (6) ◽  
pp. 1475-1483 ◽  
Author(s):  
P Vandenberg ◽  
A Kern ◽  
A Ries ◽  
L Luckenbill-Edds ◽  
K Mann ◽  
...  
Keyword(s):  
Binding Site ◽  
Type Iv Collagen ◽  
Cell Attachment ◽  
Cell Surface Receptor ◽  
Helical Conformation ◽  
Amino Acid Residues ◽  
Cell Binding ◽  
Type Iv ◽  
Depth Study ◽  
Surface Receptor

The aim of this investigation was to identify the domains of type IV collagen participating in cell binding and the cell surface receptor involved. A major cell binding site was found in the trimeric cyanogen bromide-derived fragment CB3, located 100 nm away from the NH2 terminus of the molecule, in which the triple-helical conformation is stabilized by interchain disulfide bridges. Cell attachment assays with type IV collagen and CB3 revealed comparable cell binding activities. Antibodies against CB3 inhibited attachment on fragment CB3 completely and on type IV collagen to 80%. The ability to bind cells was strictly conformation dependent. Four trypsin derived fragments of CB3 allowed a closer investigation of the binding site. The smallest, fully active triple-helical fragment was (150)3-amino acid residues long. It contained segments of 27 and 37 residues, respectively, at the NH2 and COOH terminus, which proved to be essential for cell binding. By affinity chromatography on Sepharose-immobilized CB3, two receptor molecules of the integrin family, alpha 1 beta 1 and alpha 2 beta 1, were isolated. Their subunits were identified by sequencing the NH2 termini or by immunoblotting. The availability of fragment CB3 will allow for a more in-depth study of the molecular interaction of a short, well defined triple-helical ligand with collagen receptors alpha 1 beta 1 and alpha 2 beta 1.

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