An optimized chemical synthesis of human relaxin‐2

Abstract

Human gene 2 relaxin (RLX) is a member of the insulin superfamily and is a multi-functional factor playing a vital role in pregnancy, aging, fibrosis, cardioprotection, vasodilation, inflammation, and angiogenesis. RLX is currently applied in clinical trials to cure among others acute heart failure, fibrosis, and preeclampsia. The synthesis of RLX by chemical methods is difficult because of the insolubility of its B-chain and the required laborious and low yielding site-directed combination of its A (RLXA) and B (RLXB) chains. We report here that oxidation of the Met(25) residue of RLXB improves its solubility, allowing its effective solid-phase synthesis and application in random interchain combination reactions with RLXA. Linear Met(O)(25)-RLX B-chain (RLXBO) reacts with a mixture of isomers of bicyclic A-chain (bcRLXA) giving exclusively the native interchain combination. Applying this method Met(O)(25)-RLX (RLXO) was obtained in 62% yield and was easily converted to RLX in 78% yield, by reduction with ammonium iodide.

Design and chemical syntheses of potent matriptase‐2 inhibitors based on trypsin inhibitor SFTI‐1 isolated from sunflower seeds

“…Moreover, two peptidomimetics, known as peptomers (39,40) were designed. Peptomers are peptide-peptoid hybrids, containing at least one N-substituted glycine residue (peptoid monomer).…”

“…SFTI-1 is an important and promising scaffold for drug discovery and development of pharmaceuticals. [33] were synthesized on 2-chlorotrityl chloride resin (GL Biochem, Shanghai Ltd.) and the C-terminal amino acid residue Fmoc-Asp(OtBu) was attached to the aforementioned resin as described previously [33,40] (in the presence of an equimolar amount of DIPEA under anhydrous conditions in DCM solution). Herein we describe a series of new analogues of SFTI-1 designed to be matriptase-2 inhibitors.…”

Amino Acid-Protecting Groups

“…It has been shown that the same peptide with all the hydroxyl groups protected by Trt or t Bu is obtained with better purity in the case of the former. 428 -tert-Butyldimethylsilyl (TBDMS). 424 It is more acid-labile than the t Bu group and can be removed selectively in the presence this group using AcOH-THF-H 2 O (3:1:1) or TBAF.…”

“…Thus, they are a better alternative to t Bu for the synthesis of peptides containing residues prone to alkylation such as Trp and Met. 423,441,428 Removal is carried out with 2% TFA in DCM 294 -tert-Butyldimethylsilyl (TBDMS). 424 Unlike the t Bu ethers, the TBDMS ether of Tyr is more acid-labile than the corresponding t Bu ethers; however, it can be removed selectively with TBAF.…”

Synthetic relaxins

“…It is also a potentially cheaper approach for the scale-up preparation of native synthetic protein compared to the more laborious sequential selective disulfide bond formation approach (see below). Recently, Barlos and colleagues showed that oxidation to Met(O) of one of the two Met residues within the H2 relaxin B-chain (Met25) significantly improved both its chemical synthesis and its subsequent solubility [15]. This latter aspect, in turn, enabled enhanced oxidative folding with either S-reduced or mixed isomers of disulfide bicyclic A-chain to generate correctly folded Met(O)B25-relaxin that was reduced to MetB25 with ammonium iodide in excellent overall yield (48% relative to starting B-chain).…”