Dataset Information

Simple Strategy of Integrating the Micro-centrifugation-based Extraction and 4-Hexylphenylazosulfonate for the Preparation of Plant Microsomal Proteins for Bottom-Up Proteomics

ABSTRACT: The preparation of high-quality microsomal proteins (MPs) is of critical importance to high-throughput microsomal proteomics. The isolation of MPs for proteomic analysis has to date relied on Ultracentrifugation-based methods (UBMs), however, these methods are labor-intensive, time-consuming, and unaffordable to many laboratories. Here, we report a simple method developed by integrating the micro-centrifugation-based extraction (MME) method and 4-hexylphenylazosulfonate (Azo) surfactant for the isolation and solubilization of MPs from rice leaves. A mass spectrometry-based (MS) label-free proteomic approach was used for a comprehensive quantitative comparison of the extractability between the MME method and the plasma membrane extraction (PME) kit and of the ability to solubilize proteins between Azo and sodium dodecyl sulfate (SDS). Results showed that the MME method can not only effectively isolate MPs from rice leaves for a bottom-up proteomic analysis but also performs better than the PME kit in the enrichment of plasma membrane (PM) proteins. Additionally, Azo was found to effectively solubilize total proteins, including the MPs from rice leaves for bottom-up proteomics with performance comparable to that of SDS. Azo can be integrated with the MME approach as a simple and effective strategy to prepare MPs and could replace SDS for high-throughput proteomic studies. This is the first study revealing the applicability of the micro-centrifugation-based method and Azo in plant proteomics

INSTRUMENT(S): Q Exactive

ORGANISM(S): Oryza Sativa Subsp. Japonica (rice)

TISSUE(S): Leaf

SUBMITTER: Sun Tae Kim

LAB HEAD: Sun Tae Kim

PROVIDER: PXD025132 | Pride | 2021-09-29

REPOSITORIES: Pride

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An Integrated Approach for the Efficient Extraction and Solubilization of Rice Microsomal Membrane Proteins for High-Throughput Proteomics.

Nguyen Truong Van TV Gupta Ravi R Annas Dicky D Yoon Jinmi J Kim Yu-Jin YJ Lee Gi Hyun GH Jang Jeong Woo JW Park Kang Hyun KH Rakwal Randeep R Jung Ki-Hong KH Min Cheol Woo CW Kim Sun Tae ST

Frontiers in plant science 20210909

The preparation of microsomal membrane proteins (MPs) is critically important to microsomal proteomics. To date most research studies have utilized an ultracentrifugation-based approach for the isolation and solubilization of plant MPs. However, these approaches are labor-intensive, time-consuming, and unaffordable in certain cases. Furthermore, the use of sodium dodecyl sulfate (SDS) and its removal prior to a mass spectrometry (MS) analysis through multiple washing steps result in the loss of ...[more]

PMID: 34567038

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			Action	DRS
	_Set_1__Kit_Rep_1.raw	Raw
	_Set_1__Kit_Rep_2.raw	Raw
	_Set_1__Kit_Rep_3.raw	Raw
	_Set_1__MME_Rep_1.raw	Raw
	_Set_1__MME_Rep_2.raw	Raw