-
Something wrong with this record ?
Selective chromogenic and fluorogenic peptide substrates for the assay of cysteine peptidases in complex mixtures
TA. Semashko, EA. Vorotnikova, VF. Sharikova, KS. Vinokurov, YA. Smirnova, YE. Dunaevsky, MA. Belozersky, B. Oppert, EN. Elpidina, IY. Filippova,
Language English Country United States
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Cysteine Proteases isolation & purification metabolism MeSH
- Enzyme Assays methods MeSH
- Fluorescent Dyes analysis metabolism MeSH
- Hydrolysis MeSH
- Models, Molecular MeSH
- Peptides chemistry metabolism MeSH
- Substrate Specificity MeSH
- Tenebrio enzymology metabolism MeSH
- Animals MeSH
- Check Tag
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
This study describes the design, synthesis, and use of selective peptide substrates for cysteine peptidases of the C1 papain family, important in many biological processes. The structure of the newly synthesized substrates is Glp-Xaa-Ala-Y (where Glp=pyroglutamyl; Xaa=Phe or Val; and Y=pNA [p-nitroanilide], AMC [4-amino-7-methylcoumaride], or AFC [4-amino-7-trifluoromethyl-coumaride]). Substrates were synthesized enzymatically to guarantee selectivity of the reaction and optical purity of the target compounds, simplifying the scheme of synthesis and isolation of products. The hydrolysis of the synthesized substrates was evaluated by C1 cysteine peptidases from different organisms and with different functions, including plant enzymes papain, bromelain, ficin, and mammalian lysosomal cathepsins B and L. The new substrates were selective for C1 cysteine peptidases and were not hydrolyzed by serine, aspartic, or metallo peptidases. We demonstrated an application of the selectivity of the synthesized substrates during the chromatographic separation of a multicomponent set of digestive peptidases from a beetle, Tenebrio molitor. Used in combination with the cysteine peptidase inhibitor E-64, these substrates were able to differentiate cysteine peptidases from peptidases of other classes in midgut extracts from T. molitor larvae and larvae of the genus Tribolium; thus, they are useful in the analysis of complex mixtures containing peptidases from different classes.
A N Belozersky Institute of Physico Chemical Biology Moscow State University Moscow 119992 Russia
Department of Chemistry Moscow State University Moscow 119992 Russia
Entomological Institute Biology Centre AV ĈR Ĉeské Budĕjovice CZ 37005 Czech Republic
Faculty of Bioengineering and Bioinformatics Moscow State University Moscow 119992 Russia
References provided by Crossref.org
- 000
- 00000naa a2200000 a 4500
- 001
- bmc15008419
- 003
- CZ-PrNML
- 005
- 20150310123252.0
- 007
- ta
- 008
- 150306s2014 xxu f 000 0|eng||
- 009
- AR
- 024 7_
- $a 10.1016/j.ab.2013.12.032 $2 doi
- 035 __
- $a (PubMed)24388866
- 040 __
- $a ABA008 $b cze $d ABA008 $e AACR2
- 041 0_
- $a eng
- 044 __
- $a xxu
- 100 1_
- $a Semashko, Tatiana A $u Department of Chemistry, Moscow State University, Moscow 119992, Russia.
- 245 10
- $a Selective chromogenic and fluorogenic peptide substrates for the assay of cysteine peptidases in complex mixtures / $c TA. Semashko, EA. Vorotnikova, VF. Sharikova, KS. Vinokurov, YA. Smirnova, YE. Dunaevsky, MA. Belozersky, B. Oppert, EN. Elpidina, IY. Filippova,
- 520 9_
- $a This study describes the design, synthesis, and use of selective peptide substrates for cysteine peptidases of the C1 papain family, important in many biological processes. The structure of the newly synthesized substrates is Glp-Xaa-Ala-Y (where Glp=pyroglutamyl; Xaa=Phe or Val; and Y=pNA [p-nitroanilide], AMC [4-amino-7-methylcoumaride], or AFC [4-amino-7-trifluoromethyl-coumaride]). Substrates were synthesized enzymatically to guarantee selectivity of the reaction and optical purity of the target compounds, simplifying the scheme of synthesis and isolation of products. The hydrolysis of the synthesized substrates was evaluated by C1 cysteine peptidases from different organisms and with different functions, including plant enzymes papain, bromelain, ficin, and mammalian lysosomal cathepsins B and L. The new substrates were selective for C1 cysteine peptidases and were not hydrolyzed by serine, aspartic, or metallo peptidases. We demonstrated an application of the selectivity of the synthesized substrates during the chromatographic separation of a multicomponent set of digestive peptidases from a beetle, Tenebrio molitor. Used in combination with the cysteine peptidase inhibitor E-64, these substrates were able to differentiate cysteine peptidases from peptidases of other classes in midgut extracts from T. molitor larvae and larvae of the genus Tribolium; thus, they are useful in the analysis of complex mixtures containing peptidases from different classes.
- 650 _2
- $a zvířata $7 D000818
- 650 _2
- $a cysteinové proteasy $x izolace a purifikace $x metabolismus $7 D057056
- 650 _2
- $a enzymatické testy $x metody $7 D057075
- 650 _2
- $a fluorescenční barviva $x analýza $x metabolismus $7 D005456
- 650 _2
- $a hydrolýza $7 D006868
- 650 _2
- $a molekulární modely $7 D008958
- 650 _2
- $a peptidy $x chemie $x metabolismus $7 D010455
- 650 _2
- $a substrátová specifita $7 D013379
- 650 _2
- $a Tenebrio $x enzymologie $x metabolismus $7 D013712
- 655 _2
- $a časopisecké články $7 D016428
- 655 _2
- $a práce podpořená grantem $7 D013485
- 700 1_
- $a Vorotnikova, Elena A $u Faculty of Bioengineering and Bioinformatics, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Sharikova, Valeriya F $u Department of Chemistry, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Vinokurov, Konstantin S $u Entomological Institute, Biology Centre AV ĈR, Ĉeské Budĕjovice CZ-37005, Czech Republic.
- 700 1_
- $a Smirnova, Yulia A $u A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Dunaevsky, Yakov E $u A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Belozersky, Mikhail A $u A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Oppert, Brenda $u USDA Agricultural Research Service, Center for Grain and Animal Health Research, Manhattan, KS 66502, USA. Electronic address: brenda.oppert@ars.usda.gov.
- 700 1_
- $a Elpidina, Elena N $u A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russia.
- 700 1_
- $a Filippova, Irina Y $u Department of Chemistry, Moscow State University, Moscow 119992, Russia.
- 773 0_
- $w MED00000335 $t Analytical biochemistry $x 1096-0309 $g Roč. 449, č. - (2014), s. 179-87
- 856 41
- $u https://pubmed.ncbi.nlm.nih.gov/24388866 $y Pubmed
- 910 __
- $a ABA008 $b sig $c sign $y a $z 0
- 990 __
- $a 20150306 $b ABA008
- 991 __
- $a 20150310123529 $b ABA008
- 999 __
- $a ok $b bmc $g 1065692 $s 891219
- BAS __
- $a 3
- BAS __
- $a PreBMC
- BMC __
- $a 2014 $b 449 $c - $d 179-87 $i 1096-0309 $m Analytical biochemistry $n Anal Biochem $x MED00000335
- LZP __
- $a Pubmed-20150306
