Limits...
Mapping the active site of CD59.

Yu J, Abagyan R, Dong S, Gilbert A, Nussenzweig V, Tomlinson S - J. Exp. Med. (1997)

Bottom Line: CD59 is a widely distributed membrane-bound inhibitor of the cytolytic membrane attack complex (MAC) of complement.We present data obtained from a combination of molecular modeling and mutagenesis techniques, which together indicate that the active site of CD59 is located in the vicinity of a hydrophobic groove on the face of the molecule opposite to a "hydrophobic strip" suggested earlier.In addition, removal of the single N-linked glycosylation site at Asn18 of CD59 resulted in an enhancement of complement inhibitory activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, New York University Medical Center, New York 10016, USA.

ABSTRACT
CD59 is a widely distributed membrane-bound inhibitor of the cytolytic membrane attack complex (MAC) of complement. This small (77 amino acid) glycoprotein is a member of the Ly6 superfamily of proteins and is important in protecting host cells from the lytic and proinflammatory activity of the MAC. CD59 functions by binding to C8 and/or C9 in the nascent MAC and interfering with C9 membrane insertion and polymerization. We present data obtained from a combination of molecular modeling and mutagenesis techniques, which together indicate that the active site of CD59 is located in the vicinity of a hydrophobic groove on the face of the molecule opposite to a "hydrophobic strip" suggested earlier. In addition, removal of the single N-linked glycosylation site at Asn18 of CD59 resulted in an enhancement of complement inhibitory activity.

Show MeSH

Related in: MedlinePlus

Complement resistance of CHO cells expressing CD59 and  CD59-Ly6E chimeric proteins. Complement resistance of CHO cell  populations expressing similar levels of either CD59-Ly6E or Ly6ECD59 chimeras was compared to cells expressing similar levels of CD59.  Cells were exposed to 10% NHS. Depending on whether the chimeric  protein contained CD59 at its NH2 or COOH terminus, different methods were used to quantitate relative expression. Therefore activity of chimeras should be compared to CD59 activity within each set.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196154&req=5

Figure 4: Complement resistance of CHO cells expressing CD59 and CD59-Ly6E chimeric proteins. Complement resistance of CHO cell populations expressing similar levels of either CD59-Ly6E or Ly6ECD59 chimeras was compared to cells expressing similar levels of CD59. Cells were exposed to 10% NHS. Depending on whether the chimeric protein contained CD59 at its NH2 or COOH terminus, different methods were used to quantitate relative expression. Therefore activity of chimeras should be compared to CD59 activity within each set.

Mentions: CHO cell populations expressing similar levels of recombinant CD59 or chimeric proteins were assayed for their susceptibility to human complement. The functional data obtained with the CD-Ly chimeras (CD59 at NH2 terminus) revealed that the region of CD59 which lies COOHterminal to residue 57 is not required for function; both CD-Ly 57 and CD-Ly 64 possessed the same specific activity as wild type CD59 (Fig. 4). The other CD-Ly chimeras which contained shorter CD59-specific sequences had no complement inhibitory activity. Of the Ly-CD chimeras (Ly6E at NH2 terminus), only Ly-CD 16 possessed any functional activity (Fig. 4). The specific activity of Ly-CD 16 was ∼85% that of wild-type CD59. Taken together, these data indicate that the functionally important region of CD59 is located between amino acids 16 and 57 in the primary structure.


Mapping the active site of CD59.

Yu J, Abagyan R, Dong S, Gilbert A, Nussenzweig V, Tomlinson S - J. Exp. Med. (1997)

Complement resistance of CHO cells expressing CD59 and  CD59-Ly6E chimeric proteins. Complement resistance of CHO cell  populations expressing similar levels of either CD59-Ly6E or Ly6ECD59 chimeras was compared to cells expressing similar levels of CD59.  Cells were exposed to 10% NHS. Depending on whether the chimeric  protein contained CD59 at its NH2 or COOH terminus, different methods were used to quantitate relative expression. Therefore activity of chimeras should be compared to CD59 activity within each set.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196154&req=5

Figure 4: Complement resistance of CHO cells expressing CD59 and CD59-Ly6E chimeric proteins. Complement resistance of CHO cell populations expressing similar levels of either CD59-Ly6E or Ly6ECD59 chimeras was compared to cells expressing similar levels of CD59. Cells were exposed to 10% NHS. Depending on whether the chimeric protein contained CD59 at its NH2 or COOH terminus, different methods were used to quantitate relative expression. Therefore activity of chimeras should be compared to CD59 activity within each set.
Mentions: CHO cell populations expressing similar levels of recombinant CD59 or chimeric proteins were assayed for their susceptibility to human complement. The functional data obtained with the CD-Ly chimeras (CD59 at NH2 terminus) revealed that the region of CD59 which lies COOHterminal to residue 57 is not required for function; both CD-Ly 57 and CD-Ly 64 possessed the same specific activity as wild type CD59 (Fig. 4). The other CD-Ly chimeras which contained shorter CD59-specific sequences had no complement inhibitory activity. Of the Ly-CD chimeras (Ly6E at NH2 terminus), only Ly-CD 16 possessed any functional activity (Fig. 4). The specific activity of Ly-CD 16 was ∼85% that of wild-type CD59. Taken together, these data indicate that the functionally important region of CD59 is located between amino acids 16 and 57 in the primary structure.

Bottom Line: CD59 is a widely distributed membrane-bound inhibitor of the cytolytic membrane attack complex (MAC) of complement.We present data obtained from a combination of molecular modeling and mutagenesis techniques, which together indicate that the active site of CD59 is located in the vicinity of a hydrophobic groove on the face of the molecule opposite to a "hydrophobic strip" suggested earlier.In addition, removal of the single N-linked glycosylation site at Asn18 of CD59 resulted in an enhancement of complement inhibitory activity.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, New York University Medical Center, New York 10016, USA.

ABSTRACT
CD59 is a widely distributed membrane-bound inhibitor of the cytolytic membrane attack complex (MAC) of complement. This small (77 amino acid) glycoprotein is a member of the Ly6 superfamily of proteins and is important in protecting host cells from the lytic and proinflammatory activity of the MAC. CD59 functions by binding to C8 and/or C9 in the nascent MAC and interfering with C9 membrane insertion and polymerization. We present data obtained from a combination of molecular modeling and mutagenesis techniques, which together indicate that the active site of CD59 is located in the vicinity of a hydrophobic groove on the face of the molecule opposite to a "hydrophobic strip" suggested earlier. In addition, removal of the single N-linked glycosylation site at Asn18 of CD59 resulted in an enhancement of complement inhibitory activity.

Show MeSH
Related in: MedlinePlus