An efficient method for protein extraction from low protein concentration solutions compatible with the proteomic analysis of latex-producing organelles

ABSTRACT: The extraction of high-purity proteins from the washing solution (WS) of rubber tree latex-producing organelles (also termed rubber particles) in laticifers for proteomic analysis is challenging due to the low concentration of proteins in the WS. Recent studies have revealed that proteins in the WS might play crucial roles in natural rubber biosynthesis. To further examine the involvement of these proteins in natural rubber biosynthesis, we designed an efficiency method to extract high-purity WS proteins. We improved our current borax and phenol-based (BPP) method by adding re-extraction steps with phenol (REP) to improve the yield from low-protein concentration samples. With this new method, we extracted WS proteins that were suitable for proteomics. Indeed, compared to the original BPP method, the REP method improved both the quality and quantity of isolated proteins. By repeatedly extracting from low-protein concentration solutions using the same small amount of phenol, the REP method yielded enough protein of sufficiently high-quality from starting samples containing less than 0.02 mg of proteins per mL. This method was successfully applied to extract the rubber particle proteins from the WS of natural rubber latex samples. The REP-extracted WS proteins were resolved by two-dimensional gel electrophoresis (2-DE), and 28 proteins were positively identified by mass spectrometry. This method has the potential to become widely used for the extraction of proteins from low-protein-concentration solutions for proteomic analysis.

INSTRUMENT(S): 4800 Proteomics Analyzer

ORGANISM(S): Viridiplantae

TISSUE(S): Latex

SUBMITTER: dan wang  

LAB HEAD: Xuchu Wang

PROVIDER: PXD004156 | Pride | 2016-10-10

REPOSITORIES: Pride