Publications de Kai Johnsson | EPFL Graph Search

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Genetically encoded fluorescent sensor proteins offer the possibility to probe the concentration of key metabolites in living cells. The approaches currently used to generate Such fluorescent sensor proteins lack. generality, as they require a protein that ...

A general approach for the sequential labeling of fusion proteins of O6-alkylguanine-DNA alkyltransferase (AGT) with different fluorophores in mammalian cells is presented. AGT fusion proteins with different localizations in the cell can be labeled specifically with different fluorophores, and the fluorescence labeling can be used for applications such as multicolor anal. of dynamic processes and fluorescence resonance energy transfer measurements. The facile access to a variety of different AGT substrates

Engineered nanobody allows reversible control of activity in cells through the binding of small molecules. ...

High-resolution detection of specific molecules in cells is a major challenge in biology. We show that infrared scanning near-field microscopy can detect the spatial distribution of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid–type glutamate r ...

Rhodamine dyes conjugated to targeting ligands can yield exceptionally bright fluorescent probes for live-cell imaging. However, the limited permeability of such rhodamine derivatives restricts their broader applications, particularly in vivo. Here, we present Rho-tag and SiR-tag, engineered protein tags derived from bacterial multidrug-resistant proteins that bind unsubstituted (silicon) rhodamines with nanomolar affinity. Unsubstituted (silicon) rhodamines readily cross membranes and enable rapid, reversi

A review. New, improved phenotypes. Somatic hypermutation in immune cells can be used for the fast directed evolution of proteins other than Igs (e.g., autofluorescent proteins). The target gene is inserted into the genome of activated B lymphocytes where ...

The invention pertains to a near-infrared fluorescent dye that is cell permeable and can be attached to selected proteins in living cells. The dye has the general formula (I) or its or its corresponding spirolactone (II) wherein Y is chosen from the group ...

Polo-like kinase (Plk4) is a serine/threonine-proteinkinase thatis essential for biogenesis of the centriole organelle and is enrichedat centrioles. Herein, we introduce Cen-TCO, a chemical probe basedon the Plk4 inhibitor centrinone, to image Plk4 and cen ...

Recordings of the physiological history of cells provide insights into biological processes, yet obtaining such recordings is a challenge. To address this, we introduce a method to record transient cellular events for later analysis. We designed proteins t ...

Autofluorescent proteins have become indispensable in our quest to visualize molecular events in living cells. Further progress in the visualization and quantification of all biochemical activities of the cell will require the introduction of additional an ...

We present a fluorescence activation-coupled protein labeling (FAPL) method, which employs small-molecular probes that exhibit almost no basal fluorescence but acquire strong fluorescence upon covalent binding to tag-proteins. This method enables real-time ...

Though the effect of the recently identified mitochondrial NAD(+) transporter SLC25A51 on glucose metabolism has been described, its contribution to other NAD(+)-dependent processes throughout the cell such as ADP-ribosylation remains elusive. Here, we rep ...

Wild-type catalase-peroxidase KatG from Mycobacterium tuberculosis as well as a specific mutant (R463L) frequently found in isoniazid-resistant strains have been overexpressed in Escherichia coli, allowing purifn. of sufficient quantities of enzyme for phys. and kinetic characterization. Optical absorption and EPR spectroscopies indicate that KatG is similar to a growing class of bacterial catalase-peroxidases. Optical and EPR spectra of KatG in the presence of either a strong field or weak field ligand sug

The in vivo and in vitro labeling of fusion proteins with synthetic mols. capable of probing and controlling protein function has the potential to become an important method in functional genomics and proteomics. We have recently introduced an approach for the specific labeling of fusion proteins, which is based on the generation of fusion proteins with the human DNA repair protein O6-alkylguanine-DNA alkyltransferase (hAGT) and the irreversible reaction of hAGT with O6-benzylguanine derivs. Here, we report

The role of calcium in signal transduction relies on the precise spatial and temporal control of its concentration. The existing means to detect fluctuations in Ca2+ concentrations with adequate temporal and spatial resolution are limited. We introduce her ...

Photoactivatable fluorophores are important for single-particle tracking and super-resolution microscopy. Here we present a photoactivatable fluorophore that forms a bright silicon rhodamine derivative through a light-dependent protonation. In contrast to ...

One of the most prominent self-labeling tags is SNAP-tag. It is an in vitro evolution product of the human DNA repair protein O6 -alkylguanine-DNA alkyltransferase (hAGT) that reacts specifically with benzylguanine (BG) and benzylchloropyrimidine (CP) derivatives, leading to covalent labeling of SNAP-tag with a synthetic probe (Gronemeyer et al., Protein Eng Des Sel 19:309–316, 2006; Curr Opin Biotechnol 16:453–458, 2005; Keppler et al., Nat Biotechnol 21:86–89, 2003; Proc Natl Acad Sci U S A 101:9955– 9959

Numerous synthetic fluorophores have been developed that can switch their spectroscopic properties upon interaction with other molecules or by irradiation with light. In recent years, protein-labeling techniques have been introduced that permit the specific attachment of such molecules to proteins of interest in living cells. We review here how the attachment of switchable fluorophores to selected proteins of interest via self-labeling protein tags enables new applications in different areas of biology and

Here we present a far-red, silicon-rhodamine-based fluorophore (SiR700) for live-cell multicolor imaging. SiR700 has excitation and emission maxima at 690 nm and 715 nm, respectively. SiR700-based probes for F-actin, microtubules, lysosomes and SNAP-tag ar ...

We demonstrate how a combination of self-labeling protein tags and unnatural amino acid technology permits the semisynthesis of ratiometric fluorescent sensor proteins with unprecedented dynamic range in vitro and on live cells. To generate such a sensor, a binding protein is labeled with a fluorescent competitor of the analyte using SNAP-tag in conjugation with a second fluorophore that is positioned in vicinity of the binding site of the binding protein using unnatural amino acid technology. Binding of th

Kai Johnsson

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Semisynthetic fluorescent sensor proteins based on self-labeling protein tags

Labeling of fusion proteins with synthetic fluorophores in live cells

Chemogenetic Control of Nanobodies

Photobleaching-free infrared near-field microscopy localizes molecules in neurons

Fast, Bright, and Reversible Fluorescent Labeling of Rhodamine-Binding Proteins

Fast directed evolution of non-immunoglobulin proteins by somatic hypermutation in immune cells

Cell permeable, fluorescent dye

Chemical Probe for Imaging of Polo-like Kinase 4 and Centrioles

Recording physiological history of cells with chemical labeling

Visualizing biochemical activities in living cells

Real-Time Measurements of Protein Dynamics Using Fluorescence Activation-Coupled Protein Labeling Method

Absence of mitochondrial SLC25A51 enhances PARP1-dependent DNA repair by increasing nuclear NAD(+) levels

Overexpression, purification, and characterization of the catalase-peroxidase KatG from Mycobacterium tuberculosis

Labeling of fusion proteins of O6-alkylguanine-DNA alkyltransferase with small molecules in vivo and in vitro

Indo-1 derivatives for local calcium sensing

Photoactivation of silicon rhodamines via a light-induced protonation

Fluorescent Labeling of SNAP-Tagged Proteins in Cells

Switchable fluorophores for protein labeling in living cells

Fluorogenic Probes for Multicolor Imaging in Living Cells

A General Strategy for the Semisynthesis of Ratiometric Fluorescent Sensor Proteins with Increased Dynamic Range