Facile synthesis of insulin fusion derivatives through sortase A ligation.

Disotuar, Maria M; Smith, Jake A; Li, Jinze; Alam, Steve; Lin, Nai-Pin; Chou, Danny Hung-Chieh

Disotuar, Maria M; Smith, Jake A; Li, Jinze; Alam, Steve; Lin, Nai-Pin; Chou, Danny Hung-Chieh.

Afiliación

Disotuar MM; Department of Biochemistry, School of Medicine, University of Utah, Salt Lake City, UT 84112, USA.
Smith JA; Department of Biochemistry, School of Medicine, University of Utah, Salt Lake City, UT 84112, USA.
Li J; Department of Biochemistry, School of Medicine, University of Utah, Salt Lake City, UT 84112, USA.
Alam S; Department of Biochemistry, School of Medicine, University of Utah, Salt Lake City, UT 84112, USA.
Lin NP; Department of Biochemistry, School of Medicine, University of Utah, Salt Lake City, UT 84112, USA.
Chou DH; Division of Endocrinology and Diabetes, Department of Pediatrics, School of Medicine, Stanford University, Palo Alto, CA 94305, USA.

Acta Pharm Sin B ; 11(9): 2719-2725, 2021 Sep.

Article en En | MEDLINE | ID: mdl-34589392

RESUMEN

Insulin derivatives such as insulin detemir and insulin degludec are U.S. Food and Drug Administration (FDA)-approved long-acting insulin currently used by millions of people with diabetes. These derivatives are modified in C-terminal B29 lysine to retain insulin bioactivity. New and efficient methods for facile synthesis of insulin derivatives may lead to new discovery of therapeutic insulin. Herein, we report a new method using sortase A (SrtA)-mediated ligation for the synthesis of insulin derivatives with high efficiency and functional group tolerance in the C-terminal B chain. This new insulin molecule (Ins-SA) with an SrtA-recognizing motif can be conjugated to diverse groups with N-terminal oligoglycines to generate new insulin derivatives. We further demonstrated that a new insulin derivative synthesized by this SrtA-mediated ligation shows strong cellular and in vivo bioactivity. This enzymatic method can therefore be used for future insulin design and development.

Palabras clave

Alb, albumin; Albumin-binding peptide SA21; Boc, tert-butyloxycarbonyl; DCM, dichloromethane; DIEA, N,N-diisopropylethylamine; DMEM, Dulbecco's Modified Eagle Medium; DMF, dimethylformamide; DMSO, dimethyl sulfoxide; DOI, desoctapeptide (B23−30) insulin; Diabetes mellitus; EDT, 1,2-ethanedithiol; FBS, fetal bovine serum; Fmoc, 9-fluorenylmethoxycarbonyl; HATU, 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate; HBTU, O-(benxontriazol-1-yl)-1,1,3,3-tetramethyluronium; HPLC, high performance liquid chromatography; HTRF, homogeneous time resolved fluorescence; IR-B, human insulin receptor isoform B; ITT, insulin tolerance test; Insulin synthesis; LCâMS, liquid chromatography mass spectrometry; Long-acting insulin; Mtt, 4-methyltrityl; NBD-X, 6-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)hexanoic acid; STZ, streptozotocin; Sortase A (SrtA) ligation; SrtA, sortase A; THF, triflouroacetic acid; TIS, triisoproylsilane; i.p., intraperitoneal; pAkt, phosphorylated protein kinase B; t-Bu, tert-butyl

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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Acta Pharm Sin B Año: 2021 Tipo del documento: Article País de afiliación: Estados Unidos Pais de publicación: Países Bajos

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