Limits...
The kinetics of the hydrogen/deuterium exchange of epidermal growth factor receptor ligands.

Iloro I, Narváez D, Guillén N, Camacho CM, Guillén L, Cora E, Pastrana-Ríos B - Biophys. J. (2008)

Bottom Line: All ligands were found to have similar contributions of 3(10)-helix and random coil with varying contributions of beta-sheets and beta-turns.The time constants for AR 0.47 min(-1) (Tyr), 0.04 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (buried 3(10)-helix, beta-turns, and beta-sheets); for HB-EGF 0.89 min(-1) (Tyr), 0.14 min(-1) (Arg and 3(10)-helix), and 1.00 x 10(-3) min(-1) (buried 3(10)-helix, beta-sheets, and beta-turns); and for epiregulin 0.16 min(-1) (Tyr), 0.03 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (3(10)-helix and beta-sheets).These results provide essential information toward understanding secondary structure, H/D exchange kinetics, and solvation of these epidermal growth factor receptor ligands in their unbound state.

View Article: PubMed Central - PubMed

Affiliation: Center for Protein Structure Function and Dynamics, University of Puerto Rico, Mayagüez Campus, Mayagüez, Puerto Rico.

ABSTRACT
Five highly homologous epidermal growth factor receptor ligands were studied by mass spectral analysis, hydrogen/deuterium (H/D) exchange via attenuated total reflectance Fourier transform-infrared spectroscopy, and two-dimensional correlation analysis. These studies were performed to determine the order of events during the exchange process, the extent of H/D exchange, and associated kinetics of exchange for a comparative analysis of these ligands. Furthermore, the secondary structure composition of amphiregulin (AR) and heparin-binding-epidermal growth factor (HB-EGF) was determined. All ligands were found to have similar contributions of 3(10)-helix and random coil with varying contributions of beta-sheets and beta-turns. The extent of exchange was 40%, 65%, 55%, 65%, and 98% for EGF, transforming growth factor-alpha (TGF-alpha), AR, HB-EGF, and epiregulin (ER), respectively. The rate constants were determined and classified as fast, intermediate, and slow: for EGF the 0.20 min(-1) (Tyr), 0.09 min(-1) (Arg, beta-turns), and 1.88 x 10(-3) min(-1) (beta-sheets and 3(10)-helix); and for TGF-alpha 0.91 min(-1) (Tyr), 0.27 min(-1) (Arg, beta-turns), and 1.41 x 10(-4) min(-1) (beta-sheets). The time constants for AR 0.47 min(-1) (Tyr), 0.04 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (buried 3(10)-helix, beta-turns, and beta-sheets); for HB-EGF 0.89 min(-1) (Tyr), 0.14 min(-1) (Arg and 3(10)-helix), and 1.00 x 10(-3) min(-1) (buried 3(10)-helix, beta-sheets, and beta-turns); and for epiregulin 0.16 min(-1) (Tyr), 0.03 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (3(10)-helix and beta-sheets). These results provide essential information toward understanding secondary structure, H/D exchange kinetics, and solvation of these epidermal growth factor receptor ligands in their unbound state.

Show MeSH

Related in: MedlinePlus

Overlaid difference spectra of the EGFR ligands. (A) EGF, (B) TGF-α, (C) AR, (D) HB- EGF, and (E) ER within the spectral region 1725–1400 cm−1. The difference spectra were generated by subtraction of the first spectrum from all subsequent spectra.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2367206&req=5

fig4: Overlaid difference spectra of the EGFR ligands. (A) EGF, (B) TGF-α, (C) AR, (D) HB- EGF, and (E) ER within the spectral region 1725–1400 cm−1. The difference spectra were generated by subtraction of the first spectrum from all subsequent spectra.

Mentions: The difference spectra were obtained by subtracting the first spectrum from all subsequent spectra for the spectral region of 1720–1400 cm−1. A summary of the peak assignment for the maxima (+) or minima (−) resulting from the increase or decrease in overall peak intensities is listed below. For EGF, the peak assignments are composed of β-turn ((−) 1690 and 1680 cm−1), random coil ((+) 1659 cm−1), strong β-sheet contribution ((+) 1630 cm−1), Arg side-chain modes ((+) 1600 cm−1 and 1580 cm−1), N-H deformation mode ((−) 1545 cm−1), Tyr side-chain mode ((−) 1520 cm−1), and N-D deformation mode ((+) 1450 cm−1) as shown in Fig. 4 A. For the resulting difference spectra for TGF-α difference spectra, the peak assignments are composed of β-turn ((−) 1690 cm−1), random coil ((+) 1673 cm−1), 310-helix ((+) 1646 cm−1), β-sheet ((+) 1635 cm−1), Arg side-chain modes ((+) 1600 cm−1), N-H deformation mode ((−) 1550 cm−1), Tyr side-chain mode ((−) 1520 cm−1), and N-D deformation mode ((+) 1450 cm−1), as shown in Fig. 4 B.


The kinetics of the hydrogen/deuterium exchange of epidermal growth factor receptor ligands.

Iloro I, Narváez D, Guillén N, Camacho CM, Guillén L, Cora E, Pastrana-Ríos B - Biophys. J. (2008)

Overlaid difference spectra of the EGFR ligands. (A) EGF, (B) TGF-α, (C) AR, (D) HB- EGF, and (E) ER within the spectral region 1725–1400 cm−1. The difference spectra were generated by subtraction of the first spectrum from all subsequent spectra.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2367206&req=5

fig4: Overlaid difference spectra of the EGFR ligands. (A) EGF, (B) TGF-α, (C) AR, (D) HB- EGF, and (E) ER within the spectral region 1725–1400 cm−1. The difference spectra were generated by subtraction of the first spectrum from all subsequent spectra.
Mentions: The difference spectra were obtained by subtracting the first spectrum from all subsequent spectra for the spectral region of 1720–1400 cm−1. A summary of the peak assignment for the maxima (+) or minima (−) resulting from the increase or decrease in overall peak intensities is listed below. For EGF, the peak assignments are composed of β-turn ((−) 1690 and 1680 cm−1), random coil ((+) 1659 cm−1), strong β-sheet contribution ((+) 1630 cm−1), Arg side-chain modes ((+) 1600 cm−1 and 1580 cm−1), N-H deformation mode ((−) 1545 cm−1), Tyr side-chain mode ((−) 1520 cm−1), and N-D deformation mode ((+) 1450 cm−1) as shown in Fig. 4 A. For the resulting difference spectra for TGF-α difference spectra, the peak assignments are composed of β-turn ((−) 1690 cm−1), random coil ((+) 1673 cm−1), 310-helix ((+) 1646 cm−1), β-sheet ((+) 1635 cm−1), Arg side-chain modes ((+) 1600 cm−1), N-H deformation mode ((−) 1550 cm−1), Tyr side-chain mode ((−) 1520 cm−1), and N-D deformation mode ((+) 1450 cm−1), as shown in Fig. 4 B.

Bottom Line: All ligands were found to have similar contributions of 3(10)-helix and random coil with varying contributions of beta-sheets and beta-turns.The time constants for AR 0.47 min(-1) (Tyr), 0.04 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (buried 3(10)-helix, beta-turns, and beta-sheets); for HB-EGF 0.89 min(-1) (Tyr), 0.14 min(-1) (Arg and 3(10)-helix), and 1.00 x 10(-3) min(-1) (buried 3(10)-helix, beta-sheets, and beta-turns); and for epiregulin 0.16 min(-1) (Tyr), 0.03 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (3(10)-helix and beta-sheets).These results provide essential information toward understanding secondary structure, H/D exchange kinetics, and solvation of these epidermal growth factor receptor ligands in their unbound state.

View Article: PubMed Central - PubMed

Affiliation: Center for Protein Structure Function and Dynamics, University of Puerto Rico, Mayagüez Campus, Mayagüez, Puerto Rico.

ABSTRACT
Five highly homologous epidermal growth factor receptor ligands were studied by mass spectral analysis, hydrogen/deuterium (H/D) exchange via attenuated total reflectance Fourier transform-infrared spectroscopy, and two-dimensional correlation analysis. These studies were performed to determine the order of events during the exchange process, the extent of H/D exchange, and associated kinetics of exchange for a comparative analysis of these ligands. Furthermore, the secondary structure composition of amphiregulin (AR) and heparin-binding-epidermal growth factor (HB-EGF) was determined. All ligands were found to have similar contributions of 3(10)-helix and random coil with varying contributions of beta-sheets and beta-turns. The extent of exchange was 40%, 65%, 55%, 65%, and 98% for EGF, transforming growth factor-alpha (TGF-alpha), AR, HB-EGF, and epiregulin (ER), respectively. The rate constants were determined and classified as fast, intermediate, and slow: for EGF the 0.20 min(-1) (Tyr), 0.09 min(-1) (Arg, beta-turns), and 1.88 x 10(-3) min(-1) (beta-sheets and 3(10)-helix); and for TGF-alpha 0.91 min(-1) (Tyr), 0.27 min(-1) (Arg, beta-turns), and 1.41 x 10(-4) min(-1) (beta-sheets). The time constants for AR 0.47 min(-1) (Tyr), 0.04 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (buried 3(10)-helix, beta-turns, and beta-sheets); for HB-EGF 0.89 min(-1) (Tyr), 0.14 min(-1) (Arg and 3(10)-helix), and 1.00 x 10(-3) min(-1) (buried 3(10)-helix, beta-sheets, and beta-turns); and for epiregulin 0.16 min(-1) (Tyr), 0.03 min(-1) (Arg), and 1.00 x 10(-4) min(-1) (3(10)-helix and beta-sheets). These results provide essential information toward understanding secondary structure, H/D exchange kinetics, and solvation of these epidermal growth factor receptor ligands in their unbound state.

Show MeSH
Related in: MedlinePlus