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. 2013 Apr 24;135(16):6014-7.
doi: 10.1021/ja402187t. Epub 2013 Apr 15.

Hydroxyproline-free single composition ABC collagen heterotrimer

Abhishek A Jalan  1 , Borries Demeler, Jeffrey D Hartgerink
Affiliations

Hydroxyproline-free single composition ABC collagen heterotrimer

Abhishek A Jalan et al. J Am Chem Soc. .

Abstract

Hydroxyproline plays a major role in stabilizing collagenous domains in eukaryotic organisms. Lack of this modification is associated with significant lowering in the thermal stability of the collagen triple helix and may also affect fibrillogenesis and folding of the peptide chains. In contrast, even though bacterial collagens lack hydroxyproline, their thermal stability is comparable to that of fibrillar collagen. This has been attributed to the high frequency of charged amino acids found in bacterial collagen. Here we report a thermally stable hydroxyproline-free ABC heterotrimeric collagen mimetic system composed of decapositive and decanegative peptides and a zwitterionic peptide. None of the peptides contain hydroxyproline, and furthermore the zwitterionic peptide does not even contain proline. The heterotrimer is electrostatically stabilized via multiple interpeptide lysine-aspartate and lysine-glutamate salt bridges and maintains good thermal stability with a melting temperature of 37 °C. The ternary peptide mixture also populates a single composition ABC heterotrimer as confirmed by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. This system illustrates the power of axial salt bridges to direct and stabilize the self-assembly of a triple helix and may be useful in analogous designs in expression systems where the incorporation of hydroxyproline is challenging.

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Figures

Figure 1
Figure 1
(A) Schematic illustration of hydroxyproline-free ABC heterotrimer composed of (PKG)10, (DKG)10 and (EPG)10 peptides electrostatically stabilized through multiple lysine-aspartate and lysine-glutamate salt-bridges. All three peptides contain an N-terminal YG label to assist in determination of concentration. (B) Thermal melting profile of the individual, binary and ternary mixtures of (PKG)10, (DKG)10 and (EPG)10 peptide. The monomers as well as binary mixture were monitored at 225.0 nm and the ternary mixture was monitored at 222.4 nm.
Figure 2
Figure 2
CD wavelength scan of (PKG)10•(DKG)10•(EOG)10 in blue and (PKG)10•(DKG)10•(EPG)10 in red recorded at 5 °C.
Figure 3
Figure 3
1H,15N-HSQC spectrum (A) and two-dimensional plane of a three-dimensional 1H,15N-NOESY-HSQC (B and C) spectrum of a mixture of (PKG)10, (DKG)10 and (EPG)10 peptides.
Figure 4
Figure 4
(A) NH-NH inter-chain NOE expected based on the crystal structure of (PPG)10 collagen triple helix (PDB code: 1K6F). (B) 1H,1H-NOESY spectrum of a mixture of (PKG)10, (EPG)10 and (DKG)10 showing the NH-NH NOE cross peaks. G(NH)-G(NH) correlations are indicated in black and red. The GDKG(NH)-GPKG(NH) correlation shown in orange/dashed could not be definitively assigned due to spectral overlap. (C) Model of the ABC heterotrimer composed of (PKG)10, (EPG)10 and (DKG)10 chains indicating observed NH-NH inter-chain correlations.
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References

    1. Persikov AV, Ramshaw JA, Kirkpatrick A, Brodsky B. Biochemistry. 2000;39:14960. - PubMed
    1. Ramshaw JA, Shah NK, Brodsky B. J. Struct. Biol. 1998;122:86. - PubMed
    1. Okuyama K. Conn. Tiss. Res. 2008;49:299. - PubMed
    1. Bella J. J. Struct. Biol. 2010;170:377. - PubMed
    1. Shoulders MD, Satyshur KA, Forest KT, Raines RT. Proc. Nat. Acad. Sci. USA. 2010;107:559. - PMC - PubMed

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