Based on the difference in charge and hydrophobicity between amidating moiety and the carboxyl group in a given buffer solution, a new method to analyze the C-terminal amidating structure and the amidating enzyme activity by using capillary electrophoresis was established.
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J Am Soc Mass Spectrom
January 2025
Department of Chemistry, Bagley Hall, Box 351700, University of Washington, Seattle, Washington 98195-1700, United States.
We report a study of internal covalent cross-linking with photolytically generated diarylnitrile imines of N-terminal arginine, lysine, and histidine residues in peptide conjugates. Conjugates in which a 4-(2-phenyltetrazol-5-yl)benzoyl group was attached to C-terminal lysine, that we call RAAA--K, KAAA--K, and HAAA--K, were ionized by electrospray and subjected to UV photodissociation (UVPD) at 213 nm. UVPD triggered loss of N and proceeded by covalent cross-linking to nitrile imine intermediates that involved the side chains of N-terminal arginine, lysine, and histidine, as well as the peptide amide groups.
View Article and Find Full Text PDFNat Commun
November 2024
Novo Nordisk Research Center Seattle, Inc., Seattle, WA, USA.
C-terminal α-amidated peptides are attractive therapeutic targets, but preparative methods to access amidated pharmaceuticals are limited both on lab and manufacturing-scale. Here we report a straightforward and scalable approach to the C-terminal α-amidation of peptides and proteins from cysteine-extended polypeptide precursors. This amidation protocol consists of three highly efficient steps: 1) selective cysteine thiol substitution with a photolabel, 2) photoinduced decarboxylative elimination and 3) enamide cleavage by simple acidolysis or inverse electron demand Diels-Alder reaction.
View Article and Find Full Text PDFJ Am Soc Mass Spectrom
January 2025
Department of Chemistry, Bagley Hall, Box 351700, University of Washington, Seattle, Washington 98195-1700, United States.
Peptide conjugates furnished with a 2,5-diaryltetrazolecarbonyl tag at the C-terminal lysine, which we call peptide--K, were found to undergo efficient cross-linking of Asp, Glu, Asn, and Gln residues to transient nitrile-imine intermediates produced by photodissociation and collision-induced dissociation (CID) of the tetrazole ring in gas-phase ions. UV photodissociation (UVPD) at 213 nm achieved cross-linking conversion yields of 37 and 61% for DAAAK--K and EAAAK--K, respectively. The yields for NAAAK--K and QAAAK--K were 29 and 57%, respectively.
View Article and Find Full Text PDFMolecules
November 2024
Department of Biotechnology, Universidad Politécnica de Pachuca, Zempoala 43830, Mexico.
The natural compounds PSK and PSP have antitumor and immunostimulant properties. These pharmacological benefits have been documented in vitro and in vivo, although there is no information in silico which describes the action mechanisms at the molecular level. In this study, the inverse docking method was used to identify the interactions of PSK and PSP with two local databases: BPAT with 66 antitumor proteins, and BPSIC with 138 surfaces and intracellular proteins.
View Article and Find Full Text PDFBioorg Med Chem
December 2024
Department of Chemistry, Lund University, Box 124, SE-221 00 Lund, Sweden. Electronic address:
Four directional and positional variants of sulfonamide-derivatized galactopyranosides were synthesized and evaluated against human galectin-1, -3, -4C (C-terminal), -7, -8N (N-terminal), -8C (C-terminal), -9N (N-terminal), and -9C (C-terminal), which revealed that one of the sulfonamide positions and directionalities (methyl 3-{4-[2-(phenylsulfonylamino)-phenyl]-triazolyl}-3-deoxy-α-d-galactopyranosides) bound with 6-15 fold higher affinity than the corresponding phenyltriazole (lacking the phenylsulfonamide moiety) for galectin-9N. Molecular dynamic simulations suggested that inhibitor adopted a conformation that is complementary to the galectin-9N binding site and where the sulfonamide moiety protrudes into an unexplored and non-conserved binding site perpendicular to and below the A-B subsite to interact with a His61 NH proton. This resulted in the discovery of galectin-9N inhibitors with unprecedented selectivity over other galectins, thus constituting valuable tools for studies of the biological functions of galectin-9.
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