Activity-based proteomics

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Fluorophosphonate-rhodamine (FP-Rhodamine) activity-based probe for profiling of the serine

hydrolase superfamily. In this probe the fluorophosphonate is the reactive group (RG) as it binds
irreversibly to the active-site serine nucleophile of serine hydrolases and the tag is rhodamine, a
fluorophore for in-gel visualization.

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

functional proteomic technology that uses chemical probes that react with
mechanistically related classes of enzymes.[1]

Contents

 1Description
 2Advantages
 3Multidimensional protein identification technology
 4See also
 5References

Description[edit]
The basic unit of ABPP is the probe, which typically consists of two elements: a
reactive group (RG, sometimes called a "warhead") and a tag. Additionally, some
probes may contain a binding group which enhances selectivity. The reactive group
usually contains a specially designed electrophile that becomes covalently-linked to
a nucleophilic residue in the active site of an active enzyme. An enzyme that
is inhibited or post-translationally modified will not react with an activity-based probe.
The tag may be either a reporter such as a fluorophore or an affinity label such
as biotin or an alkyne or azide for use with the Huisgen 1,3-dipolar
cycloaddition (also known as click chemistry).[2]
Advantages[edit]
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 hydrolases[3] and metalloproteases[4] that often
interact with endogenous inhibitors or that exist as inactive zymogens, this technique
offers a valuable advantage over traditional techniques that rely on abundance rather
than activity.

Multidimensional protein identification technology[edit]

In-gel ABPP using probes with different fluorophores in the same lane to simultaneously profile differences
in enzyme activities

In recent years ABPP has been combined with tandem mass spectrometry enabling

the identification of hundreds of active enzymes from a single sample. This
technique, known as ABPP-MudPIT (multidimensional protein identification
technology) is especially useful for profiling inhibitor selectivity as the potency of an
inhibitor can be tested against hundreds of targets simultaneously.
ABPP were first reported in the 1990s in the study of proteases. [5][6]

See also[edit]
 Mass spectrometry
 Proteomics
 Related inhibitors MAFP and DIFP
 Chemoproteomics

References[edit]
0. ^ Berger AB, Vitorino PM, Bogyo M (2004). "Activity-based protein profiling: applications
to biomarker discovery, in vivo imaging and drug discovery". American Journal of
Pharmacogenomics.  4 (6): 371–81. doi:10.2165/00129785-200404060-
00004. PMID 15651898.  S2CID 18637390.
1. ^ Speers AE, Adam GC, Cravatt BF (April 2003). "Activity-based protein profiling in vivo
using a copper(i)-catalyzed azide-alkyne [3 + 2] cycloaddition". Journal of the American
Chemical Society. 125 (16): 4686–7. doi:10.1021/ja034490h. PMID 12696868.
2. ^ Liu Y, Patricelli MP, Cravatt BF (December 1999).  "Activity-based protein profiling: the
serine hydrolases".  Proceedings of the National Academy of Sciences of the United
States of America. 96 (26): 14694–
 9. Bibcode:1999PNAS...9614694L. doi:10.1073/pnas.96.26.14694. PMC  24710.  PMID 
10611275.
3. ^ Saghatelian A, Jessani N, Joseph A, Humphrey M, Cravatt BF (July 2004). "Activity-
based probes for the proteomic profiling of metalloproteases".  Proceedings of the
National Academy of Sciences of the United States of America. 101 (27): 10000–
5. Bibcode:2004PNAS..10110000S. doi:10.1073/pnas.0402784101. PMC  454150.  PMI
D 15220480.
4. ^ Kam CM, Abuelyaman AS, Li Z, Hudig D, Powers JC (1993). "Biotinylated
isocoumarins, new inhibitors and reagents for detection, localization, and isolation of
serine proteases". Bioconjugate Chemistry. 4  (6): 560–
7. doi:10.1021/bc00024a021. PMID 8305526.
5. ^ Abuelyaman AS, Hudig D, Woodard SL, Powers JC (1994). "Fluorescent derivatives of
diphenyl [1-(N-peptidylamino)alkyl]phosphonate esters: synthesis and use in the
inhibition and cellular localization of serine proteases".  Bioconjugate Chemistry.  5 (5):
400–5. doi:10.1021/bc00029a004. PMID 7849068