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{{Chembox
{{Chembox
| Name = Phenylmethanesulfonyl fluoride (PMSF)
| verifiedrevid = 443307656
| verifiedrevid = 443307656
|ImageFile=PMSF.svg
| ImageFile = PMSF.svg
|ImageSize=
| ImageSize = 180
| ImageFile1 = PMSF-3D-balls.png
|IUPACName=phenylmethanesulfonyl fluoride
| ImageAlt1 = PMSF molecule
⚫
|OtherNames=
| PIN = Phenylmethanesulfonyl fluoride
⚫
|Section1= {{Chembox Identifiers
⚫
| OtherNames =
⚫
| ChemSpiderID_Ref = {{chemspidercite|correct|chemspider}}
⚫
|Section1={{Chembox Identifiers
⚫
| ChemSpiderID_Ref = {{chemspidercite|correct|chemspider}}
| ChemSpiderID = 4620
| ChemSpiderID = 4620
| KEGG_Ref = {{keggcite|correct|kegg}}
| KEGG_Ref = {{keggcite|correct|kegg}}
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| StdInChIKey = YBYRMVIVWMBXKQ-UHFFFAOYSA-N
| StdInChIKey = YBYRMVIVWMBXKQ-UHFFFAOYSA-N
| CASNo_Ref = {{cascite|correct|CAS}}
| CASNo_Ref = {{cascite|correct|CAS}}
| CASNo=329-98-6
| CASNo = 329-98-6
⚫
| UNII_Ref = {{fdacite|correct|FDA}}
⚫
| PubChem=4784
| UNII = 57KD15003I
⚫
| ChEBI_Ref = {{ebicite|correct|EBI}}
⚫
| PubChem = 4784
| ChEBI_Ref = {{ebicite|correct|EBI}}
| ChEBI = 8102
| ChEBI = 8102
| SMILES = O=S(F)(=O)Cc1ccccc1
| SMILES = O=S(F)(=O)Cc1ccccc1
| MeSHName=Phenylmethylsulfonyl+fluoride
| MeSHName = Phenylmethylsulfonyl+fluoride
}}
}}
|Section2= {{Chembox Properties
|Section2={{Chembox Properties
| C=7 | H=7 | F=1 | O=2 | S=1
| Formula=C<sub>7</sub>H<sub>7</sub>FO<sub>2</sub>S
| Appearance=Powder
| MolarMass=174.194
| Appearance=
| Density=
| Density=
| MeltingPt=
| MeltingPt=
| BoilingPt=
| BoilingPt=
| Solubility=
| Solubility=
}}
}}
|Section3= {{Chembox Hazards
|Section3={{Chembox Hazards
| MainHazards=
| MainHazards=
| FlashPt=
| FlashPt=
| AutoignitionPt =
| Autoignition=
}}
}}
}}
}}


In [[biochemistry]], '''PMSF''' (phenylmethanesulfonylfluoride or phenylmethylsulfonyl fluoride) is a serine protease inhibitor commonly used in the preparation of [[cell lysate]]s. PMSF does not inhibit all [[serine protease]]s{{Citation needed|date=January 2011}}. It is rapidly degraded in water and stock solutions are usually made up in anhydrous [[ethanol]], [[isopropanol]], [[corn oil]], or [[Dimethyl sulfoxide|DMSO]]. Proteolytic inhibition occurs when a concentration between 0.1 - 1 mM PMSF is used. The half-life is short in aqueous solutions (110 min at pH 7, 55 min at pH 7.5, and 35 min at pH 8, all at 25°C).<ref>{{cite journal | author = GT James | journal = Analytical Biochemistry | volume = 86 | issue = 2 | pages = 574 | year = 1978 | title = Inactivation of the protease inhibitor phenylmethyldulfonyl fluoride in buffers | pmid = 26289 | doi = 10.1016/0003-2697(78)90784-4}}</ref>
In [[biochemistry]], '''phenylmethylsulfonyl fluoride''' ('''PMSF''') is a [[serine protease]] inhibitor (serine hydrolase inactivator) commonly used in the preparation of [[cell lysate]]s. PMSF does not inactivate all [[serine protease]]s.<ref>{{Cite journal|url=https://www.jbc.org/article/S0021-9258(19)66050-3/fulltext|doi = 10.1074/jbc.M002758200|doi-access = free|title = Structural Basis for the Insensitivity of a Serine Enzyme (Palmitoyl-Protein Thioesterase) to Phenylmethylsulfonyl Fluoride|year = 2000|last1 = Das|first1 = Amit K.|last2 = Bellizzi|first2 = John J.|last3 = Tandel|first3 = Sagun|last4 = Biehl|first4 = Edward|last5 = Clardy|first5 = Jon|last6 = Hofmann|first6 = Sandra L.|journal = Journal of Biological Chemistry|volume = 275|issue = 31|pages = 23847–23851|pmid = 10801859}}</ref> The effective concentration of PMSF is between 0.1 - 1 mM. The half-life is short in aqueous solutions (110 min at pH 7, 55 min at pH 7.5, and 35 min at pH 8, all at 25&nbsp;°C).<ref name="James1978">{{cite journal | author = GT James | journal = Analytical Biochemistry | volume = 86 | issue = 2 | pages = 574–9 | year = 1978 | title = Inactivation of the protease inhibitor phenylmethylsulfonyl fluoride in buffers | pmid = 26289 | doi = 10.1016/0003-2697(78)90784-4}}</ref> At 4˚C, pH 8, PMSF is almost completely degraded after 1 day.<ref name="James1978"></ref> Stock solutions are usually made up in anhydrous [[ethanol]], [[isopropanol]], or [[corn oil]] and diluted immediately before use.


PMSF binds specifically to the active site [[serine]] residue in a serine protease. It does not bind to any other serine residues in the protein. This is a result of the hyperactivity of that serine residue caused by the specific environmental conditions in the enzyme's active site. Because PMSF binds covalently to the enzyme, the complex can be viewed by X-ray crystallography; it can therefore be used as a chemical label to identify an essential active site SER in an enzyme.
PMSF reacts specifically with the active site [[serine]] residue in serine hydrolases. It does not bind to any other serine residues in the protein. This is a result of the hyperactivity of that serine residue caused by the specific environmental conditions in the enzyme's active site ([[catalytic triad]]). Because PMSF bonds covalently to the enzyme, the complex can be viewed by X-ray crystallography; it can therefore be used as a chemical label to identify an essential active site serine in an enzyme.


:Enzyme(active)Ser-O-H + F-SO<sub>2</sub>CH<sub>2</sub>C<sub>6</sub>H<sub>5</sub> → EnzymeSer-O-SO<sub>2</sub>CH<sub>2</sub>C<sub>6</sub>H<sub>5</sub> + HF
:Enzyme(active)Ser-O-H + F-SO<sub>2</sub>CH<sub>2</sub>C<sub>6</sub>H<sub>5</sub> → EnzymeSer-O-SO<sub>2</sub>CH<sub>2</sub>C<sub>6</sub>H<sub>5</sub> + HF
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:Serine protease + PMSF → Irreversible enzyme-PMS complex + HF
:Serine protease + PMSF → Irreversible enzyme-PMS complex + HF


The [[median lethal dose]] between 150–215&nbsp;mg/kg<ref>{{Cite journal |last1=Myers |first1=D. K. |last2=Kemp |first2=A. |date=January 1954 |title=Inhibition of Esterases by the Fluorides of Organic Acids |url=https://www.nature.com/articles/173033a0 |journal=Nature |language=en |volume=173 |issue=4392 |pages=33–34 |doi=10.1038/173033a0 |pmid=13119739 |s2cid=4164358 |issn=1476-4687}}</ref><ref>{{Cite journal |last1=Pinsky |first1=C. |last2=Dua |first2=A. K. |last3=LaBella |first3=F. S. |date=Sep 20–27, 1982 |title=Phenylmethylsulfonyl fluoride (PMSF) given systemically produces naloxone-reversible analgesia and potentiates effects of beta-endorphin given centrally |url=https://pubmed.ncbi.nlm.nih.gov/6292607/ |journal=Life Sciences |volume=31 |issue=12–13 |pages=1193–1196 |doi=10.1016/0024-3205(82)90340-x |issn=0024-3205 |pmid=6292607}}</ref> ([[acetylcholine esterase]] inactivator). PMSF should be handled only inside a fume hood and while wearing gloves. [[Dimethyl sulfoxide|DMSO]] is sometimes recommended as solvent for stock solutions, but should not be used as it makes intact skin permeable to PMSF.
The {{LD50}} is less than 500&nbsp;mg/kg. PMSF is a [[cytotoxic]] chemical which should be handled only inside a fume hood.


==Stability==
PMSF is commonly used in protein solublization in order to deactivate proteases from digesting proteins of interest after cell lysis.
The stability of PMSF in aqueous solutions is low, as it undergoes hydrolysis with water. PMSF is reportedly stable for ~6 months at -20˚C in [[DMSO]],<ref name="Bitesize Bio 2021 m647">{{cite web | title=Protease Inhibitors 101: Best Practices for Use in the Lab | website=Bitesize Bio | date=2021-11-30 | url=https://bitesizebio.com/58195/protease-inhibitors-101/ | access-date=2023-06-22}}</ref> and 9 months at room temperature in 100% [[isopropanol]].<ref name="James1978"></ref><ref>{{cite web |title=The Complete Guide for Protease Inhibition |url=https://iti.stanford.edu/content/dam/sm/iti/documents/himc/immunoassays/ProteaseInhibitionGuide.pdf |publisher=Roche |access-date=22 June 2023}}</ref>

==Insensitive serine enzymes==
Some proteins structure limit the accessibility of comparatively bulky PMSF, and therefore PMSF is inactive against these serine enzymes like [[Palmitoyl(protein)_hydrolase|palmitoyl-protein thioesterase]].<ref name="Das2000">{{cite journal | last1=Das | first1=Amit K. | last2=Bellizzi | first2=John J. | last3=Tandel | first3=Sagun | last4=Biehl | first4=Edward | last5=Clardy | first5=Jon | last6=Hofmann | first6=Sandra L. | title=Structural Basis for the Insensitivity of a Serine Enzyme (Palmitoyl-Protein Thioesterase) to Phenylmethylsulfonyl Fluoride | journal=Journal of Biological Chemistry | publisher=Elsevier BV | volume=275 | issue=31 | year=2000 | issn=0021-9258 | doi=10.1074/jbc.m002758200 | pages=23847–23851| pmid=10801859 | doi-access=free }}</ref> Alternative sulfonyl fluoride reagents like [[p-APMSF]] and [[hexadecylsulfonylfluoride|HDSF]], have altered access to native folded protein structures, and may react with serine enzymes that PMSF cannot efficiently react with. This altered selectivity between sulfonyl fluoride reagents has been used to classify and isolate particular types of serine enzymes.<ref name="Das2000"></ref>

==See also==
* [[AEBSF]]
* [[Diisopropyl fluorophosphate|DFP]] (diisopropyl fluorophosphate), an analogous fluorophosphonate phosphorylating reagent


==References==
==References==
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{{DEFAULTSORT:Pmsf}}
{{DEFAULTSORT:Pmsf}}
[[Category:Protease inhibitors]]
[[Category:Serine protease inhibitors]]
[[Category:Serine protease inhibitors]]
[[Category:Sulfonyl halides]]
[[Category:Sulfonyl halides]]

[[de:Phenylmethylsulfonylfluorid]]
[[es:PMSF]]
[[ja:フッ化フェニルメチルスルホニル]]
[[pl:Fluorek fenylometylosulfonylu]]

Revision as of 16:45, 21 February 2024

Phenylmethanesulfonyl fluoride (PMSF)
PMSF molecule
Names
Preferred IUPAC name
Phenylmethanesulfonyl fluoride
Identifiers
3D model (JSmol)
ChEBI
ChEMBL
ChemSpider
ECHA InfoCard 100.005.774 Edit this at Wikidata
KEGG
MeSH Phenylmethylsulfonyl+fluoride
UNII
  • InChI=1S/C7H7FO2S/c8-11(9,10)6-7-4-2-1-3-5-7/h1-5H,6H2 checkY
    Key: YBYRMVIVWMBXKQ-UHFFFAOYSA-N checkY
  • InChI=1/C7H7FO2S/c8-11(9,10)6-7-4-2-1-3-5-7/h1-5H,6H2
    Key: YBYRMVIVWMBXKQ-UHFFFAOYAF
  • O=S(F)(=O)Cc1ccccc1
Properties
C7H7FO2S
Molar mass 174.19 g·mol−1
Appearance Powder
Except where otherwise noted, data are given for materials in their standard state (at 25 °C [77 °F], 100 kPa).
checkY verify (what is checkY☒N ?)

In biochemistry, phenylmethylsulfonyl fluoride (PMSF) is a serine protease inhibitor (serine hydrolase inactivator) commonly used in the preparation of cell lysates. PMSF does not inactivate all serine proteases.[1] The effective concentration of PMSF is between 0.1 - 1 mM. The half-life is short in aqueous solutions (110 min at pH 7, 55 min at pH 7.5, and 35 min at pH 8, all at 25 °C).[2] At 4˚C, pH 8, PMSF is almost completely degraded after 1 day.[2] Stock solutions are usually made up in anhydrous ethanol, isopropanol, or corn oil and diluted immediately before use.

PMSF reacts specifically with the active site serine residue in serine hydrolases. It does not bind to any other serine residues in the protein. This is a result of the hyperactivity of that serine residue caused by the specific environmental conditions in the enzyme's active site (catalytic triad). Because PMSF bonds covalently to the enzyme, the complex can be viewed by X-ray crystallography; it can therefore be used as a chemical label to identify an essential active site serine in an enzyme.

Enzyme(active)Ser-O-H + F-SO2CH2C6H5 → EnzymeSer-O-SO2CH2C6H5 + HF
Serine protease + PMSF → Irreversible enzyme-PMS complex + HF

The median lethal dose between 150–215 mg/kg[3][4] (acetylcholine esterase inactivator). PMSF should be handled only inside a fume hood and while wearing gloves. DMSO is sometimes recommended as solvent for stock solutions, but should not be used as it makes intact skin permeable to PMSF.

Stability

The stability of PMSF in aqueous solutions is low, as it undergoes hydrolysis with water. PMSF is reportedly stable for ~6 months at -20˚C in DMSO,[5] and 9 months at room temperature in 100% isopropanol.[2][6]

Insensitive serine enzymes

Some proteins structure limit the accessibility of comparatively bulky PMSF, and therefore PMSF is inactive against these serine enzymes like palmitoyl-protein thioesterase.[7] Alternative sulfonyl fluoride reagents like p-APMSF and HDSF, have altered access to native folded protein structures, and may react with serine enzymes that PMSF cannot efficiently react with. This altered selectivity between sulfonyl fluoride reagents has been used to classify and isolate particular types of serine enzymes.[7]

See also

  • AEBSF
  • DFP (diisopropyl fluorophosphate), an analogous fluorophosphonate phosphorylating reagent

References

  1. ^ Das, Amit K.; Bellizzi, John J.; Tandel, Sagun; Biehl, Edward; Clardy, Jon; Hofmann, Sandra L. (2000). "Structural Basis for the Insensitivity of a Serine Enzyme (Palmitoyl-Protein Thioesterase) to Phenylmethylsulfonyl Fluoride". Journal of Biological Chemistry. 275 (31): 23847–23851. doi:10.1074/jbc.M002758200. PMID 10801859.
  2. ^ a b c GT James (1978). "Inactivation of the protease inhibitor phenylmethylsulfonyl fluoride in buffers". Analytical Biochemistry. 86 (2): 574–9. doi:10.1016/0003-2697(78)90784-4. PMID 26289.
  3. ^ Myers, D. K.; Kemp, A. (January 1954). "Inhibition of Esterases by the Fluorides of Organic Acids". Nature. 173 (4392): 33–34. doi:10.1038/173033a0. ISSN 1476-4687. PMID 13119739. S2CID 4164358.
  4. ^ Pinsky, C.; Dua, A. K.; LaBella, F. S. (Sep 20–27, 1982). "Phenylmethylsulfonyl fluoride (PMSF) given systemically produces naloxone-reversible analgesia and potentiates effects of beta-endorphin given centrally". Life Sciences. 31 (12–13): 1193–1196. doi:10.1016/0024-3205(82)90340-x. ISSN 0024-3205. PMID 6292607.
  5. ^ "Protease Inhibitors 101: Best Practices for Use in the Lab". Bitesize Bio. 2021-11-30. Retrieved 2023-06-22.
  6. ^ "The Complete Guide for Protease Inhibition" (PDF). Roche. Retrieved 22 June 2023.
  7. ^ a b Das, Amit K.; Bellizzi, John J.; Tandel, Sagun; Biehl, Edward; Clardy, Jon; Hofmann, Sandra L. (2000). "Structural Basis for the Insensitivity of a Serine Enzyme (Palmitoyl-Protein Thioesterase) to Phenylmethylsulfonyl Fluoride". Journal of Biological Chemistry. 275 (31). Elsevier BV: 23847–23851. doi:10.1074/jbc.m002758200. ISSN 0021-9258. PMID 10801859.
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