Change search
ReferencesLink to record
Permanent link

Direct link
Covalent Protein Labeling by Enzymatic Phosphocholination
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Show others and affiliations
2015 (English)In: Angewandte Chemie International Edition, ISSN 1433-7851, E-ISSN 1521-3773, Vol. 54, no 35, 10327-10330 p.Article in journal (Refereed) Published
Abstract [en]

We present a new protein labeling method based on the covalent enzymatic phosphocholination of a specific octapeptide amino acid sequence in intact proteins. The bacterial enzyme AnkX from Legionella pneumophila has been established to transfer functional phosphocholine moieties from synthetically produced CDP-choline derivatives to N-termini, C-termini, and internal loop regions in proteins of interest. Furthermore, the covalent modification can be hydrolytically removed by the action of the Legionella enzyme Lem3. Only a short peptide sequence (eight amino acids) is required for efficient protein labeling and a small linker group (PEG-phosphocholine) is introduced to attach the conjugated cargo.

Place, publisher, year, edition, pages
2015. Vol. 54, no 35, 10327-10330 p.
Keyword [en]
enzymes, nucleotides, phosphocholination, protein modifications
National Category
Biochemistry and Molecular Biology
URN: urn:nbn:se:umu:diva-109454DOI: 10.1002/anie.201502618ISI: 000360216800050PubMedID: 26147231OAI: diva2:859709
Available from: 2015-10-08 Created: 2015-09-28 Last updated: 2016-01-27Bibliographically approved
In thesis
1. Synthesis and investigation of bacterial effector molecules
Open this publication in new window or tab >>Synthesis and investigation of bacterial effector molecules
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

During infections, bacterial microorganisms initiate profound interactions with mammalian host cells. Usually defense mechanisms of the host destroy intruding bacteria in rapid manner. However, many bacterial pathogens have evolved in a way to avoid these mechanisms. By use of effector molecules, which can be small organic molecules or proteins with enzymatic activity, the host is manipulated on a molecular level. Effectors mediating post-translational modifications (PTMs) are employed by many pathogens to influence the biological activity of host proteins. In the presented thesis, two related PTMs are investigated in detail: Adenylylation, the covalent transfer of an adenosine monophosphate group from adenosine triphosphate onto proteins, and phosphocholination, the covalent transfer of a phosphocholine moiety onto proteins. Over the past years, enzymes mediating these modifications have been discovered in several pathogens, especially as a mechanism to influence the signaling of eukaryotic cells by adenylylating or phosphocholinating small GTPases. However, the development of reliable methods for the isolation and identification of adenylylated and phosphocholinated proteins remains a vehement challenge in this field of research. This thesis presents general procedures for the synthesis of peptides carrying adenylylated or phosphocholinated tyrosine, threonine and serine residues. From the resulting peptides, mono-selective polyclonal antibodies against adenylylated tyrosine and threonine have been raised. The antibodies were used as tools for proteomic research to isolate unknown substrates of adenylyl transferases from eukaryotic cells. Mass spectrometric fragmentation techniques have been investigated to ease the identification of adenylylated proteins. Furthermore, this work presents a new strategy to identify adenylylated proteins. Additionally, small effector molecules are involved in the regulation of infection mechanisms. In this work, the small molecule LAI-1 (Legionella autoinducer 1) from the pathogen Legionella pneumophila, the causative agent of the Legionnaire’s disease, was synthesised together with its amino-derivatives. LAI-1 showed are a clear pharmacological effect on the regulation of the life cycle of L. pneumophila, initiating transmissive traits like motility and virulence. Furthermore, LAI-1 was shown to have an effect on eukaryotic cells as well. Directed motility of the eukaryotic cells was significantly reduced and the cytoskeletal architecture was reorganised, probably by interfering with the small GTPase Cdc42.

Place, publisher, year, edition, pages
Umeå: Umeå University, 2016. 110 p.
bacterial effectors, organic synthesis, Legionella, PTM, peptide synthesis, nucleotide chemistry
National Category
Organic Chemistry
Research subject
Biorganic Chemistry
urn:nbn:se:umu:diva-114698 (URN)978-91-7601-411-0 (ISBN)
Public defence
2016-02-19, KB3A9, KBC-huset, Umeå University, Umeå, 10:00 (English)
Available from: 2016-01-29 Created: 2016-01-26 Last updated: 2016-01-27Bibliographically approved

Open Access in DiVA

No full text

Other links

Publisher's full textPubMed

Search in DiVA

By author/editor
Ochtrop, PhilippAlbers, Michael F.Hedberg, Christian
By organisation
Department of Chemistry
In the same journal
Angewandte Chemie International Edition
Biochemistry and Molecular Biology

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Altmetric score

Total: 306 hits
ReferencesLink to record
Permanent link

Direct link