Penicillin G Amidase‐Catalysed Hydrolysis of Phenylacetic Hydrazides on a Solid Phase: A New Route to Enzyme‐Cleavable Linkers

Abstract

A novel catalytic property of penicillin G amidase (PGA) is described. Unexpectedly, the enzyme can hydrolyse hydrazide bonds with good efficiency, and in solution the enzyme shows a selectivity that is similar to phenylacetamides. The hydrolysis of phenylacetic hydrazides releases hydrazine, but no inhibition due to the formation of such reactive compounds was observed. This novel catalytic property was assayed also on a solid phase as a pioneering route for the design of enzyme-cleavable linkers and masked scavengers for ketones. On a solid phase a phenylacetic hydrazide compound was chemically synthesised on PEGA 1900 and PEGA þ (two co-polymers of acrylamide and ethylene glycol) and the efficiency of PGA in the release of phenylacetic acid depended on the diffusion of the protein inside the polymer. On PEGA þ the enzyme, as previously described, shows a good diffusion due to an improved electrostatic interaction with PGA thus achieving good hydrolytic conversions.

Enhancement of the enzymatic digestibility of sugarcane bagasse by steam pretreatment impregnated with hydrogen peroxide

“…Recently, the ability of PGA to hydrolyse hydrazides has been described 60. A 60% yield of hydrolysis was reported for phenylacetic hydrazides anchored on the PEGA + polymer.…”

“…This novel catalytic property was assayed on solid phase as a possible route to the design of ‘masked’ linkers, that upon selective cleavage can release the highly reactive hydrazine group 61. Upon hydrazide hydrolysis and release of the hydrazine function, an intramolecular nucleophilic attack would occur, leading to the release of the target molecule 60…”

Hydrazidase, a Novel Amidase Signature Enzyme That Hydrolyzes Acylhydrazides

“…With respect to hydrazide degradation, A. bisporus and pig kidney ␥-glutamyltransferases hydrolyze agaritine into p-(hydroxymethyl)phenylhydrazine and L-glutamate, but corresponding genes have not been identified (8,13). Some amidases and chymotrypsin hydrolyze hydrazides, but these reactions seem to be nonphysiological because the reaction rate is very low (10,(14)(15)(16)(17). Toida notably discovered an enzyme in Mycobacterium avium that hydrolyzes various hydrazides, including INH as well as nicotinoyl, benzoyl, and pyrazinoyl hydrazides, and named it hydrazidase (18).…”

New Scavenger Resin for the Reversible Linking and Monoprotection of Functionalized Aromatic Aldehydes

“…Unlike other resins, PEGA 1900 is an unique hydrophilic support and enables reactions in aqueous or other highly polar solutions. Its spacer, consisting of 43 ethylene glycol units on average and a backbone of cross-linked methyl acrylamide, reduces the loading to 0.2 mmol/g but ensures that bound molecules are fully accessible to bulky enzymes or catalysts and polar reagents that previously failed in less polar polystyrene-based PEG-ylated resins. , Several solid-phase syntheses of oligosaccharides and peptides have been reported using PEGA 1900 . On-bead screening of bound substrates is also possible …”