Matecko-Burmann, I. | Search bioRxiv

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↗

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↗