Activity based proteomics, or activity based protein profiling (ABPP) is a functional proteomic

Proteomics

Proteomics is the large-scale study of proteins, particularly their structures and functions. Proteins are vital parts of living organisms, as they are the main components of the physiological metabolic pathways of cells. The term "proteomics" was first coined in 1997 to make an analogy with...

 technology that uses specially designed chemical probes that react with mechanistically-related classes of enzymes. The basic unit of ABPP is the probe which typically consists of two elements: a reactive group (RG) and a tag. Additionally, some probes may contain a binding group which enhances selectivity. The reactive group usually contains an electrophile

Electrophile

In general electrophiles are positively charged species that are attracted to an electron rich centre. In chemistry, an electrophile is a reagent attracted to electrons that participates in a chemical reaction by accepting an electron pair in order to bond to a nucleophile...

 that gets covalently

Covalent bond

A covalent bond is a form of chemical bonding that is characterized by the sharing of pairs of electrons between atoms. The stable balance of attractive and repulsive forces between atoms when they share electrons is known as covalent bonding....

-linked to a nucleophilic

Nucleophile

A nucleophile is a species that donates an electron-pair to an electrophile to form a chemical bond in a reaction. All molecules or ions with a free pair of electrons can act as nucleophiles. Because nucleophiles donate electrons, they are by definition Lewis bases.Nucleophilic describes the...

 residue in the active site

Active site

In biology the active site is part of an enzyme where substrates bind and undergo a chemical reaction. The majority of enzymes are proteins but RNA enzymes called ribozymes also exist. The active site of an enzyme is usually found in a cleft or pocket that is lined by amino acid residues that...

 of an active enzyme

Enzyme

Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...

. An enzyme that is inhibited by enzyme inhibitor

Enzyme inhibitor

An enzyme inhibitor is a molecule that binds to enzymes and decreases their activity. Since blocking an enzyme's activity can kill a pathogen or correct a metabolic imbalance, many drugs are enzyme inhibitors. They are also used as herbicides and pesticides...

s or post-translational modifications will not react with an activity-based probe. The tag may be either a reporter such as a fluorophore

Fluorophore

A fluorophore, in analogy to a chromophore, is a component of a molecule which causes a molecule to be fluorescent. It is a functional group in a molecule which will absorb energy of a specific wavelength and re-emit energy at a different wavelength...

 or an affinity label such as biotin

Biotin

Biotin, also known as Vitamin H or Coenzyme R, is a water-soluble B-complex vitamin discovered by Bateman in 1916. It is composed of a ureido ring fused with a tetrahydrothiophene ring. A valeric acid substituent is attached to one of the carbon atoms of the tetrahydrothiophene ring...

 or an alkyne

Alkyne

Alkynes are hydrocarbons that have a triple bond between two carbon atoms, with the formula CnH2n-2. Alkynes are traditionally known as acetylenes, although the name acetylene also refers specifically to C2H2, known formally as ethyne using IUPAC nomenclature...

 or azide

Azide

Azide is the anion with the formula N3−. It is the conjugate base of hydrazoic acid. N3− is a linear anion that is isoelectronic with CO2 and N2O. Per valence bond theory, azide can be described by several resonance structures, an important one being N−=N+=N−...

 for use with the Huisgen 1,3-dipolar cycloaddition (also known as click chemistry

Click chemistry

Click chemistry is a chemical philosophy introduced by K. Barry Sharpless of The Scripps Research Institute, in 2001 and describes chemistry tailored to generate substances quickly and reliably by joining small units together...

) .

A major advantage of ABPP is the ability to monitor the availability of the enzyme active site directly, rather than being limited to protein or mRNA abundance. With classes of enzymes such as the serine protease

Serine protease

Serine proteases are enzymes that cleave peptide bonds in proteins, in which serine serves as the nucleophilic amino acid at the active site.They are found ubiquitously in both eukaryotes and prokaryotes...

s and metalloproteases that often interact with endogenous inhibitors or that exist as inactive zymogen

Zymogen

A zymogen is an inactive enzyme precursor. A zymogen requires a biochemical change for it to become an active enzyme. The biochemical change usually occurs in a lysosome where a specific part of the precursor enzyme is cleaved in order to activate it...

s, this technique offers a valuable advantage over traditional techniques that rely on abundance rather than activity.

Finally, in recent years ABPP has been combined with tandem mass spectrometry

Tandem mass spectrometry

Tandem mass spectrometry, also known as MS/MS or MS2, involves multiple steps of mass spectrometry selection, with some form of fragmentation occurring in between the stages.-Tandem MS instruments:...

 enabling the identification of hundreds of active enzymes from a single sample. This technique, known as ABPP-MudPIT is especially useful for profiling inhibitor selectivity as the potency of an inhibitor can be tested against hundreds of targets simultaneously.

ABPP was originally invented by James C. Powers (Georgia Tech, Atlanta) in the early 1990s and subsequently adopted and developed by the other labs, for example Benjamin Cravatt III

Benjamin Cravatt III

Benjamin Franklin Cravatt is a professor in and chair of the Department of Chemical Physiology at The Scripps Research Institute in La Jolla, California...

  at The Scripps Research Institute

The Scripps Research Institute

The Scripps Research Institute is an American medical research facility that focuses on research in the basic biomedical sciences. Headquartered in La Jolla, California, with a sister facility in Jupiter, Florida, the institute is home to 3,000 scientists, technicians, graduate students, and...

, Matthew Bogyo at Stanford, and others.

External links

• Description of ABPP on the Cravatt Lab website