Threonine for alanine substitution at position 109 of transthyretin differentially alters human transthyretin's affinity for iodothyronines.
The heterozygous substitution of threonine for alanine at amino acid 109 of human transthyretin (TTR) increases its affinity for T4. We compared the affinity of recombinant wild-type (WT) and Thr109-TTRs for various iodothyronines in an attempt to elucidate how this mutation alters the T4-binding site. Homozygous WT and Thr109-TTRs were expressed recombinantly in Escherichia coli, and heterozygous Thr109-TTR was purified from plasma. The affinities of the iodothyronines for TTR were determined by measuring [125I]T4 bound by TTR in the presence of increasing concentrations of unlabeled iodothyronines. Homozygous Thr109-TTR bound T4 with an affinity slightly, but not significantly, greater than that of heterozygous Thr109-TTR. The affinity of Thr109-TTR for all iodothyronines was higher than that of WT TTR. However, the Thr109 mutation increased TTR's affinity for T4, Triac (triiodothyroacetic acid), and T3 to a greater extent than it did for Tetrac (tetraiodothyroacetic acid), EMD21388 (3',5'-dibromo-4',6'-dihydroxy-3-methylflavone), and dextro-T4. These data demonstrate that a subtle change in the structure of the T4-binding channel in TTR differentially alters the affinity of binding of various iodothyronines and suggests that site-directed mutagenesis of residues within the binding channel might clarify the relative importance of specific domains of this binding channel.
Rosen
H N
HN
Diabetes Unit, Charles A. Dana Research Institute, Beth Israel Hospital, Boston, Massachusetts 02215.
Murrell
J R
JR
Liepnieks
J J
JJ
Benson
M D
MD
Cody
V
V
Moses
A C
AC
D000409:Alanine; D000595:Amino Acid Sequence; D001667:Binding, Competitive; D004926:Escherichia coli / Q000378:metabolism; D005419:Flavonoids / Q000378:metabolism; D006801:Humans; D017354:Point Mutation; D011228:Prealbumin / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins; D013237:Stereoisomerism; D013912:Threonine; D013974:Thyroxine / Q000031:analogs & derivatives* / Q000378:metabolism*; D014284:Triiodothyronine / Q000031:analogs & derivatives* / Q000378:metabolism*
D005419:Flavonoids; D011228:Prealbumin; D011994:Recombinant Proteins; D014284:Triiodothyronine; C057452:3-methyl-4',6-dihydroxy-3',5'-dibromoflavone; D013912:Threonine; D000409:Alanine; D013974:Thyroxine