Efficient Pictet–Spengler Bioconjugation with N‐Substituted Pyrrolyl Alanine Derivatives

Abstract

We discovered N-pyrrolyl alanine derivatives as efficient reagents for the fast and selective Pictet-Spengler reaction with aldehyde-containing biomolecules.O ther aldehyde-labeling methods described so far have several drawbacks,l ike hydrolytic instability,s low reaction kinetics or not readily available labeling reagents.P ictet-Spengler cyclizations of pyrrolyl 2-ethylamine substituted at the pyrrole nitrogen are significantly faster than with analogues substituted at the a-a nd b-p osition. Functionalized N-pyrrolyl alanine derivatives can be synthesized in only 2-3 steps from commercially available materials.The small sizeofthe reagent, the high reaction rate,a nd the easy synthesis make pyrrolyl alanine Pictet-Spengler (PAPS) an attractive choice for bioconjugation reactions.P APS was shown as an efficient strategy for the site-selective biotinylation of an antibody as well as for the condensation of nucleic-acid derivatives, demonstrating the versatility of this reagent.

Secondary Amino Alcohols: Traceless Cleavable Linkers for Use in Affinity Capture and Release

“…This imine is spontaneously hydrolyzed, affording a primary amine on the C‐terminal fragment ( C ) and an aldehyde on the N‐terminal fragment ( D ). Aldehydes could represent a handle for further functionalization . However, aldehydes suppress MS signals, in particular hindering MS/MS sequencing.…”

“…Aldehydes could represent ahandle for further functionalization. [26][27][28] However,a ldehydes suppress MS signals,inparticular hindering MS/MS sequencing.This issue is not only present with seramox, but also with known diol and amino alcohol linkers ( Figure 1b)a nd was circumvented by transforming the intermediate aldehydes into oximes with H 2 NOH•HCl ( Figure 2). [29] Secondary amino alcohol linkers based on isoserine (isoseramox) enable traceless release of N-terminal peptides.…”

Conversion of Serine‐Type Aldehyde Tags by the Radical SAM Protein AtsB from Methanosarcina mazei

“…This strategy is based on the conversion of a cysteine or serine residue within the conserved (C/S)X(P/A)XR motif to C α ‐formylglycine (FGly) by formylglycine‐generating enzymes (FGEs; Scheme A, B) . The aldehyde moiety of FGly can then react with specialized hydrazine or enolic nucleophiles to form hydrolytically stable protein–payload linkages (Scheme C) …”

Site-selective modification strategies in antibody–drug conjugates

“…310 A recently reported alternative strategy for formylglycine conjugation developed novel N-pyrrolyl alanine Pictet-Spengler reagents that exchanged the indole heterocycle for a pyrrole ring. 311 This approach enabled rapid antibody bioconjugation with an easily synthesised reagent. Additionally, formylglycinetagged antibodies generated by FGE have also been exploited using pyrazolone reagents, whereby ligation occurs via tandem Knoevenagel condensation-trapping reactions (Fig.…”