Peptides and proteins with unusual structure: From biophysical measurement to control of structure
Firouzbakht, Arash
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https://hdl.handle.net/2142/122097
Description
Title
Peptides and proteins with unusual structure: From biophysical measurement to control of structure
Author(s)
Firouzbakht, Arash
Issue Date
2023-12-01
Director of Research (if dissertation) or Advisor (if thesis)
Gruebele, Martin
Doctoral Committee Chair(s)
Gruebele, Martin
Committee Member(s)
Luthey Schulten, Zaida
Nair, Satish
Lu, Yi
Department of Study
Chemistry
Discipline
Chemistry
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
lasso peptides
intrinsically disordered proteins
Abstract
Despite the advancement in understanding protein structure and function, we continue to challenge our dogmas by discovering proteins and peptides with unusual structures. The harnessing of experimental, computational, and theoretical techniques has allowed researchers to carry out an integrated approach to studying these systems, from fundamental biophysical and biochemical characterization, to engineering them for improved properties and validating models. Chapter 1 introduces two classes of proteins and peptides with unusual structures, namely intrinsically disordered proteins with residual structure, and lasso peptides. The general properties of each class are described, as well as a summary of findings, in the ensuing chapters.
Chapter 2 discusses the scaling properties of marginally disordered proteins based on in vitro measurements and theoretical analysis. This work is the summary of a collaboration with Prof. Kingshuk Ghosh’s group to study the effect of environmental perturbations and charge patterning on the compaction of three proteins near the folded-unfolded boundary. We find that the target proteins vary in their disordered character and foldability and show that simple parameters such as net charge or hydropathy fail to capture the intricate scaling behavior of these disordered proteins. We reconcile the observed experimental trends with theoretical analysis and design and test variants de novo to validate the working model.
Chapter 3 investigates the dynamics of disordered protein regions in the context of chemical modifications, namely PEGylation and glycosylation. We provide an integrated experimental, computational, and theoretical study of the PEGylation and glycosylation of two variants of hirudin, partially disordered anticoagulants produced by leeches. We show that site-specific PEGylation does not perturb the thrombin-binding activity of peptide, whereas it influences the structural properties of the disordered region of unbound hirudin in a context-dependent manner. We attribute the context-dependent effect of PEG on the disordered region to specific molecular interactions that are similar but different to glycosylation.
Chapter 4 deviates from disordered proteins and focuses on the biosynthesis of lasso peptides. An in vitro reconstitution of the biosynthesis of lasso peptide fusilassin from Thermobifida fusca is elaborated. Detailed biochemical and biophysical characterization of the biosynthetic machinery unveil the molecular underpinnings of catalysis. Substrate tolerance of the enzymes are investigated, and critical catalytic residues and residues that drive the protein-protein interaction are discussed.
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