Biology subjects

Burmann, B. M.

Publications and source records attributed to Burmann, B. M..

7 recordsLinked to original sources

bioRxiv2026

Functional details of the integral membrane metallo-protease FtsH revealed by solution NMR spectroscopy

The bacterial FtsH protease resides in the inner leaflet of the bacterial inner membrane and is the sole essential protease found in Escherichia Coli (E. coli). FtsHs mandatory role is due to its proteolytic regulation of a key enzyme in the lipopolysaccharide (LPS) assembly, LpxC, although FtsH substrates comprise a diverse pool of both soluble and membrane proteins. While the full-length structure of FtsH could be solved recently, large parts of its functional cycle remain poorly understood. Here, we use advanced solution NMR spectroscopy methods in combination with biochemical assays to elucidate its ATPase cycle at an unprecedented level of detail. We reveal the foundation of the interdomain communication based on subtly tuned protein dynamics mediated via key NOE contacts as well as the kinetic rates underlying ATP consumption, providing a detailed level of insight into the interplay between the individual FtsH sub-domains, that drive the proteolytic cycle.

biophysics↗

bioRxiv2023

Disulfide-bond-induced structural frustrationand dynamic disorder in a peroxiredoxin fromMAS NMR

Disulfide bond formation is fundamentally important for protein structure, and constitutes a key mechanism by which cells regulate the intracellular oxidation state. Peroxiredoxins (PRDXs) eliminate reactive oxygen species such as hydrogen peroxide by using a catalytic cycle of Cys oxidation and reduction. High molecular-weight assemblies of PRDXs have recently been shown to additionally act as molecular chaperones. The consequences of disulfide bonds on the dynamics of these large assemblies are poorly understood. We show that formation of disulfide bonds along the catalytic cycle induces extensive s time scale dynamics, as monitored by magic-angle spinning NMR of the 216 kDa-large Tsa1 decameric assembly and solution-NMR of a designed dimeric mutant. We ascribe the conformational dynamics to structural frustration, resulting from conflicts between the disulfide-constrained reduction of mobility and the desire to fulfil other favorable contacts.

biophysics↗

bioRxiv2022

Thumb-domain dynamics modulate the functional repertoire of DNA-Polymerase IV (DinB)

In order to cope with the risk of stress-induced mutagenesis, cells in all kingdoms of life employ Y-family DNA polymerases to resolve resulting DNA lesions and thus maintaining the integrity of the genome. In Escherichia coli (E. coli) the DNA polymerase IV, or DinB, plays this crucial role in coping with these type of mutations via the so-called translesion DNA synthesis. Despite the availability of several high-resolution crystal structures important aspects of the functional repertoire of DinB remain elusive. In this study, we use advanced solution NMR spectroscopy methods in combination with biophysical characterization to elucidate the crucial role of the Thumb domain within DinBs functional cycle. We find that the inherent dynamics of this domain guide the recognition of double-stranded (ds) DNA buried within the interior of the DinB domain arrangement and trigger allosteric signals through the DinB protein. Subsequently, we characterized the RNA polymerase interaction with DinB, revealing an extended outside surface of DinB and thus not mutually excluding the DNA interaction. Altogether the obtained results lead to a refined model of the functional repertoire of DinB within the translesion DNA synthesis pathway.

biophysics↗

bioRxiv2022

Inherent dynamics of maltose binding protein (MBP) are immune to the native environment.

Biophysical characterizations of proteins typically rely on a reductionistic approach, studying proteins in a highly purified from and in absence of their natural cellular environment. Little is known about how the highly crowded conditions prevalent within living cells influence the dynamic structures proteins on the molecular level. To address this outstanding question, we characterize here the dynamic behavior of the periplasmic model protein MBP from Escherichia coli in situ, confined in the native lumen of bacterial outer membrane vesicles. To this end we determine the dynamics of side-chain methyl groups of MBP across several timescales and compare them to purified in vitro MBP. We find that the inherent dynamics of MBP are surprisingly insensitive to the native cellular environment and that the molecular motion of the protein is mainly impacted on a global level.

biophysics↗

bioRxiv2020

Bispecific tau antibodies with additional binding to C1q or alpha-synuclein

BackgroundAlzheimers disease (AD) and other tauopathies are neurodegenerative disorders characterized by cellular accumulation of aggregated tau protein. Tau pathology within these disorders is accompanied by chronic neuroinflammation, such as activation of the classical complement pathway by complement initiation factor C1q. Additionally, about half of the AD cases present with inclusions composed of aggregated alpha-synuclein called Lewy bodies. Lewy bodies in disorders such as Parkinsons disease and Lewy body dementia also frequently occur together with tau pathology. Immunotherapy is currently the most promising treatment strategy for tauopathies. However, the presence of multiple pathological processes within tauopathies makes it desirable to simultaneously target more than one disease pathway. MethodsHerein, we have developed three bispecific antibodies based on published antibody binding region sequences. One bispecific antibody binds to tau plus alpha-synuclein and two bispecific antibodies bind to tau plus C1q. ResultsThe affinity of the bispecific antibodies to their targets compared to their monospecific counterparts ranged from nearly identical to one order of magnitude lower. All bispecific antibodies retained binding to aggregated protein in patient-derived brain sections. The bispecific antibodies also retained their ability to inhibit aggregation of recombinant tau, regardless of whether the tau binding sites were in IgG or scFv format. Mono- and bispecific antibodies inhibited cellular seeding induced by AD-derived pathological tau with similar efficacy. Finally, both Tau-C1q bispecific antibodies completely inhibited the classical complement pathway. ConclusionBispecific antibodies that bind to multiple pathological targets may therefore present a promising approach to treat tauopathies and other neurodegenerative disorders.

neuroscience↗

bioRxiv2020

Structural basis of DegP-protease temperature-dependent activation

Protein quality control is an essential cellular function mainly executed by a vast array of different proteases and molecular chaperones. One of the bacterial HtrA (high temperature requirement A) protein family members, the homo-oligomeric DegP-protease, plays a crucial role in the Escherichia coli (E. coli) protein quality control machinery by removing unfolded proteins or preventing their aggregation and chaperoning them to their final folded state within the periplasm. DegP contains two regulatory PDZ domains, which play key roles in substrate recognition and in the transformation of DegP between inactive hexameric and proteolytic active cage-like structures. Here, we analyze the interaction and dynamics of the DegP PDZ-domains underlying this transformation by high-resolution NMR spectroscopy complemented with biochemical cleavage assays. We identify an interdomain molecular lock, which controls the interactions between the two PDZ domains, regulated by fine-tuned temperature-dependent protein dynamics, and which is potentially conserved in proteins harboring tandem PDZ domains.

biophysics↗

bioRxiv2020

UvrD helicase-RNA polymerase interactions are governed by UvrD's carboxy-terminal Tudor domain.

ABSTRACTAll living organisms have to cope with the constant threat of genome damage by UV light and other toxic reagents. To maintain the integrity of their genomes, organisms developed a variety of DNA repair pathways. One of these, the Transcription Coupled DNA-Repair (TCR) pathway, is triggered by stalled RNA Polymerase (RNAP) complexes at DNA damage sites on actively transcribed genes. A recently elucidated bacterial TCR pathway employs the UvrD helicase pulling back stalled RNAP complexes from the damage, stimulating recruitment of the DNA-repair machinery. However, structural and functional aspects of UvrD’s interaction with RNA Polymerase remain elusive. Here we used advanced solution NMR spectroscopy to investigate UvrD’s role within the TCR, identifying that the carboxy-terminal region of the UvrD helicase facilitates RNAP interactions by adopting a Tudor-domain like fold. Subsequently, we functionally analyzed this domain, identifying it as a crucial component for the UvrD–RNAP interaction besides having nucleic-acid affinity.Competing Interest StatementThe authors have declared no competing interest.View Full Text

biophysics↗