Fluorescent glycosidase inhibiting 1,5-dideoxy-1,5-iminoalditols

Synthesis of lipophilic 1-deoxygalactonojirimycin derivatives as D-galactosidase inhibitors

“…The isopropylidene and tert -butoxycarbonyl protecting groups were simultaneously removed under standard conditions to afford the desired free amine 15 [25], the key building block for further modifications (Scheme 2). …”

“…Different aromatic acid derivatives were prepared by coupling to the free amine at the terminus of the C 6 alkyl chain in compound 15 , which is anchored to the ring nitrogen of 1-deoxygalactonojirimycin, to yield derivatives 16 – 19 and 22 . The spacer length of six carbon units has been proven suitable for enzyme recognition in previous studies [25] and was kept constant to compare the different aromatic substituents. Additionally, Wong [15] as well as Suzuki [23] have shown from computational studies, that in case of N -substitution on compounds 8b and 10 , the iminosugar and carbasugar units respectively, were found to interact with the active site of the corresponding enzymes whereas the alkyl chains were located in the distinctly hydrophobic entrance region to the active site.…”

A Specific Activity‐Based Probe to Monitor Family GH59 Galactosylceramidase, the Enzyme Deficient in Krabbe Disease

“…Analysis was performed as previously reported. [42] Peptides were desalted on stage tips [43] and analyzed with at rap-elute system on aC 18 reversedphase nano LC with a4 5min 10-60 %ACN/0.1 %f ormic acid gradient and aT hermo LTQ-Orbitrap mass spectrometer by using a top 3d ata-dependent protocol (60 000 resolution, m/z range 300-2000, 1000 ms fill time in the Orbitrap, 35 units of CID energy for MS/MS fragmentation, 120 ms max fill time, AGC 50 e3, and 750count threshold). Ions of z = 2+ + and higher were selected to be fragmented twice within 10 s, prior to exclusion for 150 s. Peak lists were extracted and searched against the Uniprot mouse (decoy) database, with carbamidomethylation of cysteine as af ixed modification and oxidation of methionine as av ariable modification, 20 ppm peptide tolerance, trypsin as protease, and with two missed cleavages allowed by using aM ascot (matrix science) search engine.…”

“…Examples include substrates consisting of a β-galactose moiety with a luminescent or fluorogenic tag attached to the aglycon position that is released after cleavage by the enzyme, [10][11][12][13][14][15][16] and fluorescently tagged competitive inhibitors. [17,18] In addition, a few ABPs that enable non-reversible mechanism-based labeling of retaining β-galactosidases have been reported. These include suicide substrates bearing a latent quinone methide precursor as the aglycon to which a fluorescent or fluorogenic tag is attached [19][20][21][22] and 2-fluorogalactoside inhibitors in which the hydroxyl group at C6 is substituted with an azide, which enables two-step labeling via Staudinger-Bertozzi ligation.…”

“…as fluorescently tagged competitive inhibitors. [17,18] In addition, af ew ABPs that enable irreversible mechanism-basedl abeling of retaining b-galactosidases have been reported. These include suicide substrates bearing al atent quinone methide precursor as the aglycon, to whichafluorescent or fluorogenic tag is attached, [19][20][21][22] and 2-fluorogalactoside inhibitors in which the hydroxy group at C6 is substituted with an azide, which enablest wo-step labelingb yS taudinger-Bertozzi ligation.…”