Search results for "isotopic labeling"

Réponses adaptatives du Pois protéagineux à une perturbation de la fixation symbiotique d'azote en relation avec le métabolisme carboné

2013

Proteomic Analyses Reveal an Acidic Prime Side Specificity for the Astacin Metalloprotease Family Reflected by Physiological Substrates

2011

Astacins are secreted and membrane-bound metalloproteases with clear associations to many important pathological and physiological processes. Yet with only a few substrates described their biological roles are enigmatic. Moreover, the lack of knowledge of astacin cleavage site specificities hampers assay and drug development. Using PICS (proteomic identification of protease cleavage site specificity) and TAILS (terminal amine isotopic labeling of substrates) degradomics approaches >3000 cleavage sites were proteomically identified for five different astacins. Such broad coverage enables family-wide determination of specificities N- and C-terminal to the scissile peptide bond. Remarkably, me…

A Polar18F-Labeled Amino Acid Derivative for Click Labeling of Biomolecules

2014

This work describes the synthesis and 18F-labeling of an amino acid based prosthetic group that is able to participate in copper(I)-catalyzed cycloadditions. The prosthetic group can be used for 18F labeling of biomolecules under mild conditions. The synthesis started with L-serine methyl ester, which was derivatized by introducing an alkyne moiety and a leaving group for 18F labeling. Subsequently, 18F labeling as well as deprotection conditions were screened, which resulted in an overall radiochemical yield (RCY) of around 28 %. Furthermore, the 18F-labeled prosthetic group was treated with an azido cyclic Arg-Gly-Asp (cRGD) peptide as a model system in very high RCY of 98 %.

The substrate degradome of meprin metalloproteases reveals an unexpected proteolytic link between meprin β and ADAM10

2012

The in vivo roles of meprin metalloproteases in pathophysiological conditions remain elusive. Substrates define protease roles. Therefore, to identify natural substrates for human meprin α and β we employed TAILS (terminal amine isotopic labeling of substrates), a proteomics approach that enriches for N-terminal peptides of proteins and cleavage fragments. Of the 151 new extracellular substrates we identified, it was notable that ADAM10 (a disintegrin and metalloprotease domain-containing protein 10)—the constitutive α-secretase—is activated by meprin β through cleavage of the propeptide. To validate this cleavage event, we expressed recombinant proADAM10 and after preincubation with meprin…

Reverse versus Normal Prenyl Transferases in Paraherquamide Biosynthesis Exhibit Distinct Facial Selectivities

1999

Both a face-selective and a non-face-selective mode of formation of quaternary centers of isoprene-derived structural moieties of the natural alkaloid paraherquamide A (1) have been discovered by feeding experiments on Penicillium fellutanum with [U-13 C6 ]-glucose and [13 C2 ]-acetate. The labeling patterns suggest that the methyl groups (C22, C23) are introduced in a non-face-selective manner by a reverse prenyl transferase. The C5 unit comprising the dioxepin moiety retains stereochemical integrity indicative of a single, face-selective addition of the phenolic group to the dimethylallyl group.

Elucidation of the biosynthesis and degradation of allantofuranone by isotopic labelling and fermentation of modified precursors.

2010

Feeding experiments with the ascomycete Allantophomopsis lycopodina indicated that the potent fungistatic allantofuranone is biosynthesized from phenylalanine. Further experiments with synthetic precursors gave evidence that the naturally occurring polyporic acid serves as a key intermediate in the biosynthesis. In addition to the formation of allantofuranone, its abiotic and metabolic degradation were investigated.

Infrared Difference Spectroscopy of Proteins: From Bands to Bonds

2020

Infrared difference spectroscopy probes vibrational changes of proteins upon their perturbation. Compared with other spectroscopic methods, it stands out by its sensitivity to the protonation state, H-bonding, and the conformation of different groups in proteins, including the peptide backbone, amino acid side chains, internal water molecules, or cofactors. In particular, the detection of protonation and H-bonding changes in a time-resolved manner, not easily obtained by other techniques, is one of the most successful applications of IR difference spectroscopy. The present review deals with the use of perturbations designed to specifically change the protein between two (or more) functional…

Techniques for Biosynthesis

2015

A simple, rapid method for the preparation of [11C]formaldehyde **

2008

Metalloprotease meprin beta generates nontoxic N-terminal amyloid precursor protein fragments in vivo.

2011

Identification of physiologically relevant substrates is still the most challenging part in protease research for understanding the biological activity of these enzymes. The zinc-dependent metalloprotease meprin β is known to be expressed in many tissues with functions in health and disease. Here, we demonstrate unique interactions between meprin β and the amyloid precursor protein (APP). Although APP is intensively studied as a ubiquitously expressed cell surface protein, which is involved in Alzheimer disease, its precise physiological role and relevance remain elusive. Based on a novel proteomics technique termed terminal amine isotopic labeling of substrates (TAILS), APP was identified …