TY - JOUR
T1 - Mechanistically different catalytic antibodies obtained from immunization with a single transition-state analog
AU - Guo, Jincan
AU - Huang, Wei
AU - Zhou, G. Wayne
AU - Fletterick, Robert J.
AU - Scanlan, Thomas S.
PY - 1995/2/28
Y1 - 1995/2/28
N2 - The variable-region peptide sequence and steady-state kinetic behavior are compared for a family of catalytic antibodies that arose from the same immune response to a transition-state analog. The crystal structure of the most catalytically active member of the family (17E8) has been solved to 2.5 Å resolution and shows that the antibody active site contains a Ser(H99)- His(H35) (H = heavy chain) catalytic dyad analogous to the Ser-His-Asp catalytic triad of serine proteases. The variable-region peptide sequence of the next most active antibody (29G11) differs from that of 17E8 by nine heavy-chain point mutations, and results from computer modeling suggest that the three-dimensional structure of 29G11 is similar to that of 17E8. In addition, 29G11 is an efficient catalytic antibody; it possesses 26% of the hydrolytic activity of 17E8. There is one active-site mutation in 29G11 compared to 17E8; position 99 of the heavy chain of 29G11 contains a glycine residue in place of the nucleophilic serine at this position in 17E8. Consistent with this mutation, results from pH-rate studies and hydroxylamine partitioning experiments indicate that in contrast to the catalytic mechanism of 17E8, the mechanism of 29G11-catalyzed esterolysis does not feature nucleophilic catalysis.
AB - The variable-region peptide sequence and steady-state kinetic behavior are compared for a family of catalytic antibodies that arose from the same immune response to a transition-state analog. The crystal structure of the most catalytically active member of the family (17E8) has been solved to 2.5 Å resolution and shows that the antibody active site contains a Ser(H99)- His(H35) (H = heavy chain) catalytic dyad analogous to the Ser-His-Asp catalytic triad of serine proteases. The variable-region peptide sequence of the next most active antibody (29G11) differs from that of 17E8 by nine heavy-chain point mutations, and results from computer modeling suggest that the three-dimensional structure of 29G11 is similar to that of 17E8. In addition, 29G11 is an efficient catalytic antibody; it possesses 26% of the hydrolytic activity of 17E8. There is one active-site mutation in 29G11 compared to 17E8; position 99 of the heavy chain of 29G11 contains a glycine residue in place of the nucleophilic serine at this position in 17E8. Consistent with this mutation, results from pH-rate studies and hydroxylamine partitioning experiments indicate that in contrast to the catalytic mechanism of 17E8, the mechanism of 29G11-catalyzed esterolysis does not feature nucleophilic catalysis.
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U2 - 10.1073/pnas.92.5.1694
DO - 10.1073/pnas.92.5.1694
M3 - Article
C2 - 7878042
AN - SCOPUS:0028955273
SN - 0027-8424
VL - 92
SP - 1694
EP - 1698
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 5
ER -