Activity-based profiling of glycoconjugate processing enzymes
Witte, M.D.
Citation
Witte, M. D. (2009, December 22). Activity-based profiling of glycoconjugate processing enzymes. Retrieved from https://hdl.handle.net/1887/14551
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ACTIVITY-BASED PROFILING OF GLYCOCONJUGATE PROCESSING
ENZYMES
Proefschrift ter verkrijging van
de graad van Doctor aan de Universiteit Leiden, op gezag van Rector Magnificus prof. mr. P.F. van der Heijden,
volgens besluit van het College voor Promoties te verdedigen op dinsdag 22 december 2009
klokke 13.45 uur door
Martin Daudi Witte
Geboren te Bukumbi (Missungwi district te Tanzania) in 1981
Promotie commissie
Promotores : Prof. dr. H.S. Overkleeft Prof. dr. G.A. van der Marel Co-promotor : Dr. B.I. Florea
Overige leden : Prof. dr. J.M.F.G. Aerts Dr. M. Bogyo
Prof. dr. J. Brouwer Prof. dr. J. Lugtenburg
Dr. S.H.L. Verhelst
Table of contents
List of abbreviations 5
General introduction and outline 7
BODIPY-VAD-Fmk, 33 a useful tool to study yeast peptide N-glycanase activity
A chitobiose based peptide N-glycanase 55 inhibitor library, synthesis and biological evaluation
O-GlcNAc peptide epoxyketones 81 are recognized by mammalian proteasomes
Cyclophellitol based glucosidase probes 95 Synthesis and biological evaluation
1
2
3
4
5
Ultrasensitive visualization of active GBA1 111 Activity-based labeling in vitro and in situ
Summary, work in progress and future prospects 123
Samenvatting 135
List of publications 138
Curriculum vitae 140
Acknowledgements 141
6 7
List of abbreviations
AAA
ATPase ATPase associated with cellular activities
ABP activity-based probe ABPP activity-based protein profiling Ac acetyl
Ahx aminohexanoic acid APT attached proton test ATP adenosine triphosphate aq. aqueous
Az azidoacetyl Bn benzyl
Boc tert-butyloxycarbonyl
Boc2O tert-butyloxycarbonic anhydride BODIPY boron-dipyrromethene,
boradiazaindacene BOP benzotriazol-1-yl-oxy-tris-
(dimethylamino) phosphonium hexafluorophosphate BSA bovine serum albumin Bu butyl
Bz benzoyl calcd. calculated cat. catalytic amount CBB coomassie briliant blue CBE conduritol B epoxide CDI carbonyldiimidazole CMP cytidine monophosphate
chemical shift d doublet Da Dalton
DBU diazabicyclo[5.4.0]undec-7-ene DC dual color marker
DCE 1,2-dichloroethane dd double doublet ddd double double doublet DIC N,N-diisopropyl carbodiimide DiPEA diisopropylethylamine DMAP 4-(dimethylamino) pyridine DMBA 2,6-dimethylbenzoic acid DMEM dulbecco’s modified eagle’s
medium
DMF N,N-dimethylformamide DMSO dimethylsulfoxide DMT 4,4’-dimethoxytrityl DPP diphenylphosphinic acid
dt double triplet DTT dithiothreitol dq double quartet EDC 1-ethyl-3-(3-dimethyl-
aminopropyl)-carbodiimide EDEM ER degradation enhancing -
mannosidase I like proteins EDTA ethylenediaminetetraacetate ER endoplasmic reticulum ERAD ER-associated degradation
pathway Et3N triethylamine Et2O diethyl ether EtOH ethanol
EtOAc ethyl acetate equiv. molar equivalent ESI electron spray ionization
Et ethyl
Fmk fluoromethylketone Fmoc (9H-fluoren-9-yl)
methoxycarbonyl GA glycosylasparaginase Gal galactose
GalNAc N-acetylgalactosamine GBA-1 human acid -glucosidase,
glucocerebrosidase GBA-2 acid -glucosidase 2,
nonlysosomal glucocerebrosidase GBA-3 broad specifity -glucosidase Glc glucose
GlcNAc N-acetylglucosamine GDP guanosine diphosphate h. hour(s)
HCTU (2-(6-chloro-1H-benzotriazole-1- yl)-1,1,3,3-tetramethylaminium hexafluorophosphate
HEK293T human embryonic kidney cell line HMPB 4-(4-hydroxymethyl-3-
methoxyphenoxy)-butyric acid HOBt N-hydroxybenzotriazole HPLC high performance liquid
chromatography HRMS high resolution mass
spectrometry
Hz Hertz
IC50 inhibition concentration resulting in 50% inhibition of enzyme activity
ICAT isotope-coded affinity tagging IPTG isopropyl -D-
thiogalactopyranoside J coupling constant K Kelvin
LacZ E. coli -galactosidase
LC/MS liquid chromatography/mass spectrometry
m multiplet M molar
MALDI-MS matrix-assisted laser desorption/ionization mass spectrometry
Man mannose
MBHA p-methylbenzhydryl amine mCPBA meta-chloroperoxybenzoic acid MHC I major histocompatibility complex
class I Me methyl MeCN acetonitrile MeOD CD3OD MeOH methanol min. minute(s) MS molecular sieves Ms methanesulfonyl (mesyl) m/z mass-to-charge ratio NBS N-bromosuccinimide n-BuOH n-butanol
NIS N-iodosuccinimide NMM N-methylmorpholine NMO 4-methylmorpholine N-oxide NMP N-methyl-2-pyrrolidone NMR nuclear magnetic resonance PBS phosphate buffered saline Pd/C palladium on charcoal
Pd(OH)2 palladium hydroxide on charcoal PE petroleum ether
PEG polyethyleneglycol Ph phenyl
Phth phthaloyl
PNGase peptide N-glycanase ppm parts per million pTsOH para-toluenesulfonic acid
monohydrate pyr pyridine q quartet quant. quantitative RP reverse phase
rt room temperature Rt retention time s singlet sat. saturated
SDS sodium dodecyl sulfate SDS-PAGE sodium dodecyl sulfate-poly
acrylamide gel electroforesis sp. species
Su succinimidyl t triplet t, tert tertiary T temperature
TAP transporter associated with antigen presentation TBAF tetrabutylammonium fluoride TBAI tetrabutylammonium iodide TBS tert-butyldimethylsilyl TBS tris bufferd saline
TBTA tris[(1-benzyl-1H-1,2,3-triazol-4- yl)methyl] amine
td triple doublet TES triethylsilane TFA trifluoroacetic acid TFE trifluoroethanol
Tf trifluoromethanesulfonyl (triflate) TFDO methyl(trifluormethyl) dioxirane THF tetrahydrofuran
TIS triisopropyl silane TLC thin layer chromatography TMR tetramethylrhodamine TMS tetramethylsilane Tol toluene
Tris 2-amino-2-(hydroxymethyl)-1,3- propanediol
TTBP 2,4,6-tri-tert-butyl pyrimidine UDP uridine diphosphate
UGGT UDP-glucose:glycoprotein glucosyltransferase
VCNagZ Vibrio cholerae NagZ YPng1 yeast peptide N-glycanase Z benzyloxycarbonyl
