Adam Belsom

Technische Universität Berlin | TUB · Institute of Biotechnology

Chemical cross-linking combined with mass spectrometry has proven useful for studying protein-protein interactions and protein structure, however the low resolution of cross-linking has so far precluded its use in determining structures de novo. Cross-linking resolution has been typically limited by the chemical selectivity of the standard cross-li...

Hybrid methods combine experimental data and computational modeling to analyze protein structures that are elusive to structure determination. To spur the development of hybrid methods, we propose to test them in the context of the CASP experiment and would like to invite experimental groups to participate in this initiative.

Use of a heterobifunctional photoactivatable cross-linker, sulfo-SDA (diazirine), has yielded high-density data that facilitated structure modelling of individual proteins. We expand the photoactivatable chemistry toolbox here with a second reagent, sulfo-SBP (benzophenone). This further increases the density of photo-cross-linking to a factor of 2...

Highlights The earliest structural studies on proteins using crosslinking/mass spectrometry aimed to elucidate their tertiary three-dimensional structure. Tertiary structure modeling using crosslinking fell out of favor for almost two decades because crosslink data were not informative to aid structure modeling. Two game-changing trends emerged: us...

Determining the structure of a protein by any method requires various contributions from experimental and computational sides. In a recent study, high-density cross-linking/mass spectrometry (HD-CLMS) data in combination with ab initio structure prediction determined the structure of human serum albumin (HSA) domains, with an RMSD to X-ray structur...

[NiFe]-hydrogenases catalyze the reversible activation of H2 using a unique NiFe(CN)2CO metal site, which is assembled by a sophisticated multiprotein machinery. The [4Fe–4S] cluster-containing HypCD complex, which possesses an ATPase activity with a hitherto unknown function, serves as the hub for the assembly of the Fe(CN)2CO subfragment. HypCD i...

Crosslinking mass spectrometry provides pivotal information on the structure and interaction of proteins. MS-cleavable crosslinkers are regarded as a cornerstone for the analysis of complex mixtures. Yet they fragment under similar conditions as peptides, leading to mixed fragmentation spectra of the crosslinker and peptide. This hampers selecting...

Artificial metalloenzymes (ArMs) enrich bioorthogonal chemistry with new-to-nature reactions while limiting metal deactivation and toxicity. This enables biomedical applications such as activating therapeutics in situ. However, while combination therapies are becoming widespread anticancer treatments, dual catalysis by ArMs has not yet been shown....

Crosslinking mass spectrometry has become a core technology in structural biology and is expanding its reach towards systems biology. Its appeal lies in a rapid workflow, high sensitivity and the ability to provide data on proteins in complex systems, even in whole cells. The technology depends heavily on crosslinking reagents. The anatomy of cross...

Heterogeneous aggregates of the human protein α-synuclein (αSyn) are abundantly found in Lewy body inclusions of Parkinson’s disease patients. While structural information on classical αSyn amyloid fibrils is available, little is known about the conformational properties of disease-relevant, non-canonical aggregates. Here, we analyze the structural...

Heterogeneous aggregates of the human protein α-synuclein (αSyn) are abundantly found in Lewy body inclusions of Parkinson′s disease patients. While structural information on classical αSyn amyloid fibrils is available, little is known about the conformational properties of disease-relevant, non-canonical aggregates. Here, we analyze the structural...

We analyzed the backbone fragmentation behavior of tryptic peptides of a four protein mixture and of E. coli lysate subjected to Ultraviolet Photodissociation (UVPD) at 213 nm on a commercially available UVPD-equipped tribrid mass spectrometer. We obtained 15,178 unique high-confidence peptide UVPD spectrum matches by re-cording a reference beam-ty...

We analyzed the backbone fragmentation behavior of tryptic peptides of a four protein mixture and of E. coli lysate subjected to Ultraviolet Photodissociation (UVPD) at 213 nm on a commercially available UVPD-equipped tribrid mass spectrometer. We obtained 15,178 high-confidence peptide-spectrum matches by additionally recording a reference beam-ty...

Abstract We present a concise workflow to enhance the mass spectrometric detection of crosslinked peptides by introducing sequential digestion and the crosslink identification software xiSEARCH. Sequential digestion enhances peptide detection by selective shortening of long tryptic peptides. We demonstrate our simple 12‐fraction protocol for crossl...

With the advance of experimental procedures obtaining chemical crosslinking information is becoming a fast and routine practice. Information on crosslinks can greatly enhance the accuracy of protein structure modeling. Here, we review the current state of the art in modeling protein structures with the assistance of experimentally determined chemic...

Crosslinking mass spectrometry draws structural information from covalently-linked peptide pairs. When these links do not match to previous structural models, they may indicate changes in protein conformation. Unfortunately, such links can also be the result of experimental error or artefacts. Here, we describe the observation of non-covalently-ass...

Crosslinking mass spectrometry draws structural information from covalently-linked peptide pairs. When these links do not match to previous structural models, they may indicate changes in protein conformation. Unfortunately, such links can also be the result of experimental error or artefacts. Here, we describe the observation of non-covalently-ass...

We present a concise workflow to enhance the mass spectrometric detection of cross-linked peptides by introducing sequential digestion and the cross-link identification software Xi. Sequential digestion enhances peptide detection by shortening long tryptic peptides while avoiding over-digestion. We demonstrate our simple 12-fraction protocol for cr...

Experimental data offers empowering constraints for structure prediction. These constraints can be used to filter equivalently scored models or more powerfully within optimization functions toward prediction. In CASP12, Small Angle X-ray Scattering (SAXS) and Cross-Linking Mass Spectrometry (CLMS) data, measured on an exemplary set of novel fold ta...

In our paper entitled "Optimized Fragmentation Regime for Diazirine Photo-Cross-Linked Peptides", there are two separate textual errors, one in Figure 6B (and repeated in the TOC graphic) and one in the Data Analysis section. (Chemical Equation Presented) Results: Decision tree representation (Figure 6B and TOC graphic) In Figure 6B and again in th...

Little is known about how archaeal viruses perturb the transcription machinery of their hosts. Here we provide the first example of an archaeo-viral transcription factor that directly targets the host RNA polymerase (RNAP) and efficiently represses its activity. ORF145 from the temperate Acidianus two-tailed virus (ATV) forms a high-affinity comple...

Supplementary Figures 1-5 and Supplementary Table 1 and 2, Supplementary Reference.

Little is known about how archaeal viruses perturb the transcription machinery of their hosts. Here we provide the first example of an archaeo-viral transcription factor that directly targets the host RNA polymerase (RNAP) and efficiently represses its activity. ORF145 from the temperate Acidianus two-tailed virus (ATV) forms a high-affinity comple...

Cross-linking/mass spectrometry has evolved into a robust technology that reveals structural insights into proteins and protein complexes. We leverage a new tribrid instrument with improved fragmentation capacities in a systematic comparison to identify which fragmentation method would be best for the identification of cross-linked peptides. Specif...

Determining the structure of a protein by any method requires varies contributions from experimental and computational sides. In a recent study, high-density cross-linking/mass spectrometry data in combination with ab initio structure prediction by conformational space search determined the structure of human serum albumin (HSA) domains, with an RM...

Hybrid approaches combine computational methods with experimental data. The information contained in the experimental data can be leveraged to probe the structure of proteins otherwise elusive to computational methods. Compared with computational methods, the structures produced by hybrid methods exhibit some degree of experimental validation. In s...

For the first time the influence of the chirality of the gel fibers in protein crystallogenesis has been studied. Enantiomeric hydrogels and were tested with model proteins lysozyme and glucose isomerase and a formamidase extracted from B. cereus. Crystallization behaviour and crystal quality of these proteins in both hydrogels are presented and co...

FRET based systems are some of the best methods available to detect and monitor proteolytic activity. To enhance fluorescent signals and hence assay sensitivity, two different systems were developed using two different dendrimeric constructs. In the first case, a triple branched dendrimer bearing three dansyl groups was used to enhance assay sensit...