A method for whole protein isolation from human cranial bone

The presence of the dense hydroxyapatite matrix within human bone limits the applicability of conventional protocols for protein extraction. This has hindered the complete and accurate characterization of the human bone proteome thus far, leaving many bone-related disorders poorly understood. We sou...

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Bibliographic Details
Published in:Analytical biochemistry 2016-12, Vol.515, p.33-39
Main Authors: Lyon, Sarah M., Mayampurath, Anoop, Rogers, M. Rose, Wolfgeher, Donald J., Fisher, Sean M., Volchenboum, Samuel L., He, Tong-Chuan, Reid, Russell R.
Format: Article
Language:English
Subjects:
Animals
Cranial bone
Durapatite - chemistry
Extraction
Human
Humans
Method
Mice
Protein
Proteome - isolation & purification
Proteome - metabolism
Proteomics - methods
Skull - chemistry
Skull - metabolism
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Summary:The presence of the dense hydroxyapatite matrix within human bone limits the applicability of conventional protocols for protein extraction. This has hindered the complete and accurate characterization of the human bone proteome thus far, leaving many bone-related disorders poorly understood. We sought to refine an existing method of protein extraction from mouse bone to extract whole proteins of varying molecular weights from human cranial bone. Whole protein was extracted from human cranial suture by mechanically processing samples using a method that limits protein degradation by minimizing heat introduction to proteins. The presence of whole protein was confirmed by western blotting. Mass spectrometry was used to sequence peptides and identify isolated proteins. The data have been deposited to the ProteomeXchange with identifier PXD003215. Extracted proteins were characterized as both intra- and extracellular and had molecular weights ranging from 9.4 to 629 kDa. High correlation scores among suture protein spectral counts support the reproducibility of the method. Ontology analytics revealed proteins of myriad functions including mediators of metabolic processes and cell organelles. These results demonstrate a reproducible method for isolation of whole protein from human cranial bone, representing a large range of molecular weights, origins and functions.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2016.09.021