Orientation and conformation of osteocalcin adsorbed onto calcium phosphate and silica surfaces

Citation

Scudeller, L. A.; Srinivasan, S.; Rossi, A. M.; Stayton, P. S.; Drobny, G. P.; & Castner, D. G. (2017). Orientation and conformation of osteocalcin adsorbed onto calcium phosphate and silica surfaces. Biointerphases, 12(2).

Abstract

Adsorption isotherms, circular dichroism (CD) spectroscopy, x-ray photoelectron spectroscopy (XPS), and time-of-flight secondary ion mass spectrometry (ToF-SIMS) were used to investigate the adsorption of human osteocalcin (hOC) and decarboxylated (i.e., Gla converted back to Glu) hOC (dhOC) onto various calcium phosphate surfaces as well as silica surfaces. The adsorption isotherms and XPS nitrogen signals were used to track the amount of adsorbed hOC and dhOC. The intensities of key ToF-SIMS amino acid fragments were used to assess changes in the structure of adsorbed hOC and dhOC. CD spectra were used to investigate the secondary structure of OC. The largest differences were observed when the proteins were adsorbed onto silica versus calcium phosphate surfaces. Similar amounts (3-4 at. % N) of hOC and dhOC were adsorbed onto the silica surface. Higher amounts of hOC and dhOC were adsorbed on all the calcium phosphate surfaces. The ToF-SIMS data showed that the intensity of the Cys amino acid fragment, normalized to intensity of all amino acid fragments, was significantly higher (similar to x10) when the proteins were adsorbed onto silica. Since in the native OC structure the cysteines are located in the center of three a-helices, this indicates both hOC and dhOC are more denatured on the silica surface. As hOC and dhOC denature upon adsorption to the silica surface, the cysteines become more exposed and are more readily detected by ToF-SIMS. No significant differences were detected between hOC and dhOC adsorbed onto the silica surface, but small differences were observed between hOC and dhOC adsorbed onto the calcium phosphate surfaces. In the OC structure, the alpha-3 helix is located above the alpha-1 and alpha-2 helices. Small differences in the ToF-SIMS intensities from amino acid fragments characteristic of each helical unit (Asn for alpha-1; His for alpha-2; and Phe for alpha-3) suggests either slight changes in the orientation or a slight uncovering of the alpha-1 and alpha-2 for adsorbed dhOC. XPS showed that similar amounts of hOC and dhOC were absorbed onto hydroxyapaptite and octacalcium phosphate surfaces, but ToF-SIMS detected some small differences in the amino acid fragment intensities on these surfaces for adsorbed hOC and dhOC. (C) 2017 American Vacuum Society.

Keyword(s)

beta-cell
bone
calcification
gla protein
glucose-metabolism
ion mass-spectrometry
Mice
protein films
spectroscopy
tof-sims data

Notes

Ev4rh
Times Cited:0
Cited References Count:46

Reference Type

Journal Article

Secondary Title

Biointerphases

Author(s)

Scudeller, L. A.
Srinivasan, S.
Rossi, A. M.
Stayton, P. S.
Drobny, G. P.
Castner, D. G.

Year Published

2017

Date Published

1496275200

Volume Number

Issue Number

ISSN/ISBN

1934-8630

DOI

Artn 02d411
10.1116/1.4983407