Aggregate structure of hydroxyproline-rich glycoprotein (HRGP) and HRGP assisted dispersion of carbon nanotubes

Nanoscale Research Letters, Dec 2006

Hydroxyproline-rich glycoproteins (HRGP) comprise a super-family of extracellular structural glycoproteins whose precise roles in plant cell wall assembly and functioning remain to be elucidated. However, their extended structure and repetitive block co-polymer character of HRGPs may mediate their self-assembly as wall scaffolds by like-with-like alignment of their hydrophobic peptide and hydrophilic glycopeptide modules. Intermolecular crosslinking further stabilizes the scaffold. Thus the design of HRGP-based scaffolds may have practical applications in bionanotechnology and medicine. As a first step, we have used single-molecule or single-aggregate atomic force microscopy (AFM) to visualize the structure of YK20, an amphiphilic HRGP comprised entirely of 20 tandem repeats of: Ser-Hyp4-Ser-Hyp-Ser-Hyp4-Tyr-Tyr-Tyr-Lys. YK20 formed tightly aggregated coils at low ionic strength, but networks of entangled chains with a porosity of ~0.5–3 μm at higher ionic strength. As a second step we have begun to design HRGP-carbon nanotube composites. Single-walled carbon nanotubes (SWNTs) can be considered as seamless cylinders rolled up from graphene sheets. These unique all-carbon structures have extraordinary aromatic and hydrophobic properties and form aggregated bundles due to strong inter-tube van der Waals interactions. Sonicating aggregated SWNT bundles with aqueous YK20 solubilized them presumably by interaction with the repetitive, hydrophobic, Tyr-rich peptide modules of YK20 with retention of the extended polyproline-II character. This may allow YK20 to form extended structures that could potentially be used as scaffolds for site-directed assembly of nanomaterials.

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Aggregate structure of hydroxyproline-rich glycoprotein (HRGP) and HRGP assisted dispersion of carbon nanotubes

Ben Wegenhart 0 Li Tan 0 Michael Held 0 Marcia Kieliszewski 0 Liwei Chen 0 0 B. Wegenhart L. Tan M. Held M. Kieliszewski L. Chen (&) Department of Chemistry and Biochemistry, Ohio University , Athens, Ohio 45701, USA Hydroxyproline-rich glycoproteins (HRGP) comprise a super-family of extracellular structural glycoproteins whose precise roles in plant cell wall assembly and functioning remain to be elucidated. However, their extended structure and repetitive block co-polymer character of HRGPs may mediate their self-assembly as wall scaffolds by like-with-like alignment of their hydrophobic peptide and hydrophilic glycopeptide modules. Intermolecular crosslinking further stabilizes the scaffold. Thus the design of HRGP-based scaffolds may have practical applications in bionanotechnology and medicine. As a first step, we have used single-molecule or single-aggregate atomic force microscopy (AFM) to visualize the structure of YK20, an amphiphilic HRGP comprised entirely of 20 tandem repeats of: Ser-Hyp4-Ser-Hyp-Ser-Hyp4-TyrTyr-Tyr-Lys. YK20 formed tightly aggregated coils at low ionic strength, but networks of entangled chains with a porosity of ~0.5-3 lm at higher ionic strength. As a second step we have begun to design HRGPcarbon nanotube composites. Single-walled carbon nanotubes (SWNTs) can be considered as seamless cylinders rolled up from graphene sheets. These unique all-carbon structures have extraordinary aromatic and hydrophobic properties and form aggregated bundles due to strong inter-tube van der Waals interactions. Sonicating aggregated SWNT bundles with aqueous YK20 solubilized them presumably by interaction with the repetitive, hydrophobic, Tyr-rich peptide modules of YK20 with retention of the extended polyproline-II character. This may allow YK20 to form extended structures that could potentially be used as scaffolds for site-directed assembly of nanomaterials. - Published online: 1 August 2006 to the authors 2006 Hydroxyproline-rich glycoproteins (HRGPs) comprise a superfamily of extra-cellular structural proteins expressed in plant cell walls and extracellular matrix during normal development and in response to stress [1, 2]. HRGPs are extended macromolecules consisting of small repetitive peptide and glycopeptide motifs. While the peptide motifs often contain hydrophobic tyrosine residues, the glycopeptide motifs result from a combination of post-translational modifications unique to plants, namely proline hydroxylation and subsequent hydroxyproline (Hyp) glycosylation. The precise oligosaccharides or polysaccharide decoration pattern is driven by a sequence-dependent glycosylation code [24]. The key to this glycosylation code is Hyp contiguity: contiguous Hyp residues direct the addition of small arabinooligosaccharides to Hyp, while clustered non-contiguous Hyp residues direct the addition of larger complex hetero-polysaccharides. The addition of short oligosaccharides to Hyp residues locks the contiguous Hyp-rich glycopeptide motifs into an extended, left-handed polyproline-II helix conformation and thus results in rigid hydrophilic regions. In contrast, regions that lack contiguous Hyp remain flexible while subsequent addition of long polysaccharide to clustered non-contiguous Hyp residues promotes an extended random coil conformation [3]. Some HRGPs also contain hydrophobic, tyrosinerich peptide motifs that function in intra- and intermolecular crosslinking. Indeed, using a synthetic gene approach we recently expressed in tobacco cells a simple arabinosylated HRGP analog containing 20 tandem repeats of the sequence: Ser-Hyp4-Ser-HypSer-Hyp4-Tyr-Tyr-Tyr-Lys, designated YK20, and demonstrated that YK20 was extensively crosslinked enzymically in vitro to give tyrosine-based intermolecular crosslinks [5]. This indicated that YK20 rapidly aligns itself for subsequent intermolecular crosslinking and raised questions about the aggregate structure of YK20 that drives this self-assembly, the networks that arise and whether or not their properties can be tailored for specific applications. Here we report the first visualization of an YK20 network by the single-molecule or single-aggregate imaging approach using atomic force microscopy (AFM), the first such characterization for any HRGP. We also noted that YK20, an amphiphilic molecule, interacted with single-walled carbon nanotubes (SWNTs) and dispersed SWNTs in aqueous solutions, which raised the possibility that SWNT-YK20 complexes might be exploited to yield templates for the assembly of high order structures. YK20 synthetic gene construction, plant cell transformation and YK20 glycoprotein isolation A synthetic gene, YK20-EGFP, encoding 20 tandem repeats of the protein sequence Ser-Pro4-Ser-Pro-SerPro4-Tyr-Tyr-Tyr-Lys fused to the gene for the enhanced green fluorescent protein (EGFP; Clontech) was constructed, tobacco cells (Bright Yellow 2) transformed, and the YK20 glycoprotein isolated after EGFP removal, all as previously (...truncated)


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Ben Wegenhart, Li Tan, Michael Held, Marcia Kieliszewski, Liwei Chen. Aggregate structure of hydroxyproline-rich glycoprotein (HRGP) and HRGP assisted dispersion of carbon nanotubes, Nanoscale Research Letters, 2006, pp. 154, Volume 1, Issue 2, DOI: 10.1007/s11671-006-9006-8