Generation of Proteins with Free N-Terminal Cysteine by Aminopeptidases.
Journal
Journal of the American Chemical Society
ISSN: 1520-5126
Titre abrégé: J Am Chem Soc
Pays: United States
ID NLM: 7503056
Informations de publication
Date de publication:
30 11 2022
30 11 2022
Historique:
pubmed:
16
11
2022
medline:
2
12
2022
entrez:
15
11
2022
Statut:
ppublish
Résumé
Efficient, site-specific, and bio-orthogonal conjugation of chemical functionalities to proteins is of great utility in fundamental research as well as industrial processes (e.g., the production of antibody-drug conjugates and immobilization of enzymes for biocatalysis). A popular approach involves reacting a free N-terminal cysteine with a variety of electrophilic reagents. However, current methods for generating proteins with N-terminal cysteines have significant limitations. Herein we report a novel, efficient, and convenient method for producing recombinant proteins with free N-terminal cysteines by genetically fusing a Met-Pro-Cys sequence to the N-terminus of a protein of interest and subjecting the recombinant protein to the sequential action of methionine and proline aminopeptidases. The resulting protein was site-specifically labeled at the N-terminus with fluorescein and a cyclic cell-penetrating peptide through native chemical ligation and a 2-cyanobenzothiazole moiety, respectively. In addition, the optimal recognition sequence of
Identifiants
pubmed: 36378906
doi: 10.1021/jacs.2c10194
pmc: PMC9923720
mid: NIHMS1870897
doi:
Substances chimiques
Cysteine
K848JZ4886
Aminopeptidases
EC 3.4.11.-
Recombinant Proteins
0
Fluorescein
TPY09G7XIR
Peptide Library
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
21763-21771Subventions
Organisme : NCI NIH HHS
ID : P30 CA016058
Pays : United States
Organisme : NIGMS NIH HHS
ID : R01 GM110406
Pays : United States
Organisme : NIGMS NIH HHS
ID : R35 GM122459
Pays : United States
Organisme : NIH HHS
ID : S10 OD018507
Pays : United States
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