Cellulosomes are polyprotein machineries that efficiently degrade cellulosic material. Crucial to their function are scaffolds consisting of highly homologous cohesin domains, which serve a dual role by coordinating a multiplicity of enzymes as well as anchoring the microbe to its substrate. Here we combined two approaches to elucidate the mechanical properties of the main scaffold ScaA of Acetivibrio cellulolyticus. A newly developed parallelized one-pot in vitro transcription-translation and protein pull-down protocol enabled high-throughput atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) measurements of all cohesins from ScaA with a single cantilever, thus promising improved relative force comparability. Albeit very similar in sequence, the hanging cohesins showed considerably lower unfolding forces than the bridging cohesins, which are subjected to force when the microbe is anchored to its substrate. Additionally, all-atom steered molecular dynamics (SMD) simulations on homology models offered insight into the process of cohesin unfolding under force. Based on the differences among the individual force propagation pathways and their associated correlation communities, we designed mutants to tune the mechanical stability of the weakest hanging cohesin. The proposed mutants were tested in a second high-throughput AFM SMFS experiment revealing that in one case a single alanine to glycine point mutation suffices to more than double the mechanical stability. In summary, we have successfully characterized the force induced unfolding behavior of all cohesins from the scaffoldin ScaA, as well as revealed how small changes in sequence can have large effects on force resilience in cohesin domains. Our strategy provides an efficient way to test and improve the mechanical integrity of protein domains in general.
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Publication date: Available online 4 January 2018 Source: European Journal of Radiology Author(s): Peiyao Zhang, Jing Wang, Qin Xu, Zhen...
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Publication date: March 2017 Source: Free Radical Biology and Medicine, Volume 104 from #AlexandrosSfakianakis via Alexandros G.Sfak...
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Dtsch med Wochenschr DOI: 10.1055/s-0043-100054 Hintergrund und Fragestellung Ein etablierter Weg, die optimale Behandlung von Tumorpatien...
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Deepak Thapa, Vanita Ahuja, Deepanshu Dhiman Indian Journal of Anaesthesia 2017 61(12):1012-1014 from #AlexandrosSfakianakis via Alexa...
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BACKGROUND AND PURPOSE: Dengue is a common arboviral disease, which uncommonly involves the brain. There has been a recent surge in dengu...
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Abstract Limited memory size is considered as a major bottleneck in data centers for intelligent urban computing. It is shown that there e...
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Linked Article: Maintz et al. Br J Dermatol 2017; 176:481–487 . from #AlexandrosSfakianakis via Alexandros G.Sfakianakis on Inoreader h...
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Publication date: Available online 28 February 2018 Source: Oral Surgery, Oral Medicine, Oral Pathology and Oral Radiology Author(s): Mo...
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