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Identification of Protein Hydroxylation Site

The hydroxylation of proteins catalyzed by dioxygen-dependent enzymes is now considered an important post-translational modification. Hydroxylation is a chemical reaction in which a hydroxyl group (-OH) is introduced into the chemical molecule to replace the hydrogen atom covalently linked carbon. In organic chemistry, hydroxylation converts a -G-H bond into a -C-OH bond, and this chemical reaction is also known as oxidation. Thus, hydroxylation converts aliphatic hydrocarbons to aliphatic alcohols. For example, hydroxylation converts methane to methanol, ethane to ethanol, butane to butanol, etc. Similarly, organic aromatic hydrocarbons are converted to aromatic alcohols by hydroxylation, such as the hydroxylation of benzene which converts benzene to phenol.

Protein Hydroxylation

Functions of Protein Hydroxylation

Hydroxylation plays a very important role in the structure and function of organic molecules and biomolecules. It converts hydrophobic molecules into hydrophilic molecules, thus enhancing the overall solubility. In addition, hydroxylated molecules preserve better cleavage by the kidneys and liver, so they tend to excrete more easily than non-hydroxylated molecules.

To elaborate further, hydroxylation converts phenylalanine (Phe) residues to tyrosine (Tyr) residues, thus playing an important role in controlling the excess of Phe residues in the organism. Hydroxylation plays an important role in the conversion of Try to I-DOPA, which is important for the biosynthesis of dopamine. Neurotransmitters such as epinephrine are catecholamine hormones derived from dopamine. Therefore, hydroxylation is essential for the production of neurotransmitters. Nevertheless, proline hydroxylation plays an important role in the homeostasis of protein structure.

Hydroxylation Mass Spectrometry Detection Service

Hydroxylation adds an oxygen atom with a mass of 16 Da, and this increase in molecular weight can be easily detected by high-resolution mass spectrometry.

Relying on the professional HPLC-MS/MS mass spectrometry platform, Creative Proteomics can efficiently and accurately identify protein hydroxylation sites in eukaryotic and prokaryotic samples.

Service Workflow

Service Workflow

Enzymatic digestion in-gel or in-solution
Enrichment of target protein peptides
HPLC separation followed by MS/MS analysis
Hydroxylation site data analysis


Wide range of applications: characterization of hydroxylation sites for various proteins; identification of specific modification sites and specifically modified peptides

Simple and convenient operation: no radioisotope labeling or other complex pre-processing require

High site coverage (optional): enrichment of hydroxylated target proteins for increased detection coverage

Sample requirements

1. Tissue samples
Plant tissue samples: >400 mg
Blood samples: ≥2 mL (with EDTA for anticoagulation in plasma)
Serum: 2 mL
Urine: 10 mL
Animal tissue samples: ≥ 2 g
Cell samples: 1 x 10^8 cells
Yeast, microorganisms and others: dry weight 400 mg

2. Protein sample: total protein of 2-5 mg; common tissue and cell lysate can be used for protein extraction

3. Sample shipping: Please use sufficient dry ice for shipping


1. Experimental procedures
2. Mass Spectrometry Parameters
3. Details of hydroxylation sites post-translational modifications
4. Mass Spectrogram

1. Schofield C., Murray-Rust T. Enzyme Catalyzed Post-Translational Hydroxylation of Proteins. Encyclopedic Reference of Genomics and Proteomics in Molecular Medicine. Springer, (2006).

* For Research Use Only. Not for use in diagnostic procedures.
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