TY - JOUR
T1 - Comparison of native and recombinant non-phosphorylated human β-casein
T2 - Further evidence for a unique β-casein folding pattern
AU - Bu, Hongyin
AU - Hu, Yilin
AU - Sood, Satish M.
AU - Slattery, Charles W.
N1 - Recombinant wild-type non-phosphorylated human beta-casein was obtained from Escherichia coli. Turbidity vs. temperature (T) without Ca(2+) showed wild-type self-association like native except for irreversibility upon T-cycling with the original pattern re-established after concentrated urea/dialysis. With Ca(2+), wild-type was more native-like. Intrinsic Trp fluorescence spectra were similar but with lowered intensity for the wild-type protein.
PY - 2003/7/15
Y1 - 2003/7/15
N2 - Recombinant wild-type non-phosphorylated human β-casein was obtained from Escherichia coli. Turbidity vs. temperature (T) without Ca2+ showed wild-type self-association like native except for irreversibility upon T-cycling with the original pattern re-established after concentrated urea/dialysis. With Ca2+, wild-type was more native-like. Intrinsic Trp fluorescence spectra were similar but with lowered intensity for the wild-type protein. Changes in extrinsic ANS fluorescence from 4 to 37°C showed less exposure of hydrophobic surface for wild-type than native. Trp to ANS fluorescence resonance energy transfer was higher for wild-type than native at 4°C but 2- to 3-fold lower at 37°C. The native protein must be directed by the environment and/or a chaperone to fold into a unique, somewhat flexible, conformation, unaltered by urea during purification. Wild-type protein, with many native properties, does not spontaneously fold to the native conformation, even after solubilization with urea. T-cycling gives a stable conformation that is different from the native.
AB - Recombinant wild-type non-phosphorylated human β-casein was obtained from Escherichia coli. Turbidity vs. temperature (T) without Ca2+ showed wild-type self-association like native except for irreversibility upon T-cycling with the original pattern re-established after concentrated urea/dialysis. With Ca2+, wild-type was more native-like. Intrinsic Trp fluorescence spectra were similar but with lowered intensity for the wild-type protein. Changes in extrinsic ANS fluorescence from 4 to 37°C showed less exposure of hydrophobic surface for wild-type than native. Trp to ANS fluorescence resonance energy transfer was higher for wild-type than native at 4°C but 2- to 3-fold lower at 37°C. The native protein must be directed by the environment and/or a chaperone to fold into a unique, somewhat flexible, conformation, unaltered by urea during purification. Wild-type protein, with many native properties, does not spontaneously fold to the native conformation, even after solubilization with urea. T-cycling gives a stable conformation that is different from the native.
KW - Fluorescence intensity
KW - Fluorescence resonance energy transfer
KW - Human β-casein
KW - Protein folding
KW - Protein self-association
KW - Thermal cycling
KW - Turbidity
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U2 - 10.1016/S0003-9861(03)00276-5
DO - 10.1016/S0003-9861(03)00276-5
M3 - Article
C2 - 12831844
SN - 0003-9861
VL - 415
SP - 213
EP - 220
JO - Archives of Biochemistry and Biophysics
JF - Archives of Biochemistry and Biophysics
IS - 2
ER -