scholarly journals Definition of surface-exposed epitopes on the (Ca2+-Mg2+)-ATPase of sarcoplasmic reticulum

1991 ◽  
Vol 279 (1) ◽  
pp. 203-212 ◽  
Author(s):  
R E A Tunwell ◽  
J W Conlan ◽  
I Matthews ◽  
J M East ◽  
A G Lee
Keyword(s):  
Monoclonal Antibodies ◽  
Sarcoplasmic Reticulum ◽  
Polyclonal Antibodies ◽  
Nucleotide Binding ◽  
Binding Domain ◽  
Nucleotide Binding Domain ◽  
Stalk Domain ◽  
Definition Of ◽  
Peptide Antibodies

Epitopes for monoclonal antibodies binding to the native (Ca(2+)-Mg2+)-ATPase have been defined by studying binding to sets of hexameric peptides synthesized on plastic pegs. Epitopes have been confirmed by demonstrating the binding of anti-peptide antibodies to the ATPase. A method is presented for definition of surface-exposed epitopes using polyclonal antibodies. Three surface-exposed epitopes have been defined in the nucleotide-binding domain of the ATPase, suggesting considerable surface exposure of this region. Other surface-exposed epitopes have been located in the region of the fourth stalk domain.

scholarly journals Definition of surface-exposed and trans-membranous regions of the (Ca2+-Mg2+)-ATPase of sarcoplasmic reticulum using anti-peptide antibodies

1992 ◽  
Vol 286 (2) ◽  
pp. 567-580 ◽  
Author(s):  
A M Mata ◽  
I Matthews ◽  
R E A Tunwell ◽  
R P Sharma ◽  
A G Lee ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Alpha Helix ◽  
Nucleotide Binding ◽  
Proteinase K ◽  
Immunological Methods ◽  
Binding Domains ◽  
Peptide Antibody ◽  
Proteinase K Treatment ◽  
Definition Of ◽  
Peptide Antibodies

Peptides have been synthesized representing parts of the transduction, phosphorylation, nucleotide-binding and hinge domains of the (Ca(2+)-Mg2+)-ATPase of skeletal muscle sarcoplasmic reticulum (SR), and corresponding to segments of all of the postulated short inter-membranous loops of the (Ca(2+)-Mg2+)-ATPase (residues 77-88, 277-287, 780-791, 808-818, 915-924 and 949-958). A number of antibodies raised to these peptides have been shown to bind to the ATPase, defining surface-exposed regions. Many of these are concentrated in the phosphorylation and nucleotide-binding domains, suggesting that these domains could be exposed on the top surface of the ATPase. The cytoplasmic location of the loop containing residues 808-818 was confirmed by the finding that proteinase K treatment of intact SR vesicles enhanced the binding of antibodies against this segment. These findings support the 10-alpha-helix model of the ATPase. These results also suggest that only inter-membranous loops larger than about 20 residues are likely to be detected by immunological methods in transmembranous proteins. Binding of anti-peptide antibodies to proteolytic fragments of the ATPase has been used to define the domain structure of the enzyme. Some of the anti-peptide antibodies have been characterized by studying their binding to sets of hexameric peptides synthesized on plastic pegs. A wide pattern of responses is observed, with a restricted range of epitopes being recognized by each anti-peptide antibody.

scholarly journals A possible nucleotide-binding domain in the tertiary fold of phosphoribosyltransferases.

1983 ◽  
Vol 258 (10) ◽  
pp. 6450-6457 ◽  
Author(s):  
P Argos ◽  
M Hanei ◽  
J M Wilson ◽  
W N Kelley
Keyword(s):  
Nucleotide Binding ◽  
Binding Domain ◽  
Nucleotide Binding Domain
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