What Is Phosphorylation and How Does It Work?

Oxidative, Glucose, and Protein Phosphorylation

The adenosine triphosphate (ATP) molecule is formed by phosphorylation of adenine.

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Phosphorylation is the chemical addition of a phosphoryl group (PO3-) to an organic molecule. The removal of a phosphoryl group is called dephosphorylation. Both phosphorylation and dephosphorylation are carried out by enzymes (e.g., kinases, phosphotransferases). Phosphorylation is important in the fields of biochemistry and molecular biology because it's a key reaction in protein and enzyme function, sugar metabolism, and energy storage and release.

Purposes of Phosphorylation

Phosphorylation plays a critical regulatory role in cells. Its functions include:

  • Important for glycolysis
  • Used for protein-protein interaction
  • Used in protein degradation
  • Regulates enzyme inhibition
  • Maintains homeostasis by regulating energy-requiring chemical reactions

Types of Phosphorylation

Many types of molecules can undergo phosphorylation and dephosphorylation. Three of the most important types of phosphorylation are glucose phosphorylation, protein phosphorylation, and oxidative phosphorylation.

Glucose Phosphorylation

Glucose and other sugars are often phosphorylated as the first step of their catabolism. For example, the first step of glycolysis of D-glucose is its conversion into D-glucose-6-phosphate. Glucose is a small molecule that readily permeates cells. Phosphorylation forms a larger molecule that can't easily enter tissue. So, phosphorylation is critical for regulating blood glucose concentration. Glucose concentration, in turn, is directly related to glycogen formation. Glucose phosphorylation is also linked to cardiac growth.

Protein Phosphorylation

Phoebus Levene at the Rockefeller Institute for Medical Research was the first to identify a phosphorylated protein (phosvitin) in 1906, but enzymatic phosphorylation of proteins wasn't described until the 1930s.

Protein phosphorylation occurs when the phosphoryl group is added to an amino acid. Usually, the amino acid is serine, although phosphorylation also occurs on threonine and tyrosine in eukaryotes and histidine in prokaryotes. This is an esterification reaction where a phosphate group reacts with the hydroxyl (-OH) group of a serine, threonine, or tyrosine side chain. The enzyme protein kinase covalently binds a phosphate group to the amino acid. The precise mechanism differs somewhat between prokaryotes and eukaryotes. The best-studied forms of phosphorylation are posttranslational modifications (PTM), which means the proteins are phosphorylated after translation from an RNA template. The reverse reaction, dephosphorylation, is catalyzed by protein phosphatases.

An important example of protein phosphorylation is the phosphorylation of histones. In eukaryotes, DNA is associated with histone proteins to form chromatin. Histone phosphorylation modifies the structure of chromatin and alters its protein-protein and DNA-protein interactions. Usually, phosphorylation occurs when DNA is damaged, opening up space around broken DNA so that repair mechanisms can do their work.

In addition to its importance in DNA repair, protein phosphorylation plays a key role in metabolism and signaling pathways.

Oxidative Phosphorylation

Oxidative phosphorylation is how a cell stores and releases chemical energy. In a eukaryotic cell, the reactions occur within the mitochondria. Oxidative phosphorylation consists of the reactions of the electron transport chain and those of chemiosmosis. In summary, redox reaction pass electrons from proteins and other molecules along the electron transport chain in the inner membrane of the mitochondria, releasing energy that is used to make adenosine triphosphate (ATP) in chemiosmosis.

In this process, NADH and FADH2 deliver electrons to the electron transport chain. Electrons move from higher energy to lower energy as they progress along the chain, releasing energy along the way. Part of this energy goes to pumping hydrogen ions (H+) to form an electrochemical gradient. At the end of the chain, electrons are transferred to oxygen, which bond with H+ to form water. H+ ions supply the energy for ATP synthase to synthesize ATP. When ATP is dephosphorylated, cleaving the phosphate group releases energy in a form the cell can use.

Adenosine is not the only base that undergoes phosphorylation to form AMP, ADP, and ATP. For example, guanosine may also form GMP, GDP, and GTP.

Detecting Phosphorylation

Whether or not a molecule has been phosphorylated can be detected using antibodies, electrophoresis, or mass spectrometry. However, identifying and characterizing phosphorylation sites is difficult. Isotope labeling is often used, in conjunction with fluorescence, electrophoresis, and immunoassays.


  • Kresge, Nicole; Simoni, Robert D.; Hill, Robert L. (2011-01-21). "The Process of Reversible Phosphorylation: the Work of Edmond H. Fischer". Journal of Biological Chemistry. 286 (3).
  • Sharma, Saumya; Guthrie, Patrick H.; Chan, Suzanne S.; Haq, Syed; Taegtmeyer, Heinrich (2007-10-01). "Glucose Phosphorylation Is Required for Insulin-Dependent mTOR Signalling in the Heart". Cardiovascular Research. 76 (1): 71–80.
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Helmenstine, Anne Marie, Ph.D. "What Is Phosphorylation and How Does It Work?" ThoughtCo, Feb. 16, 2021, thoughtco.com/phosphorylation-definition-4140732. Helmenstine, Anne Marie, Ph.D. (2021, February 16). What Is Phosphorylation and How Does It Work? Retrieved from https://www.thoughtco.com/phosphorylation-definition-4140732 Helmenstine, Anne Marie, Ph.D. "What Is Phosphorylation and How Does It Work?" ThoughtCo. https://www.thoughtco.com/phosphorylation-definition-4140732 (accessed June 5, 2023).