Nanomaterials

Editorial

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4 pages, 215 KiB

Open AccessEditorial

Protein Nanomechanics

by Gabriel Žoldák

Nanomaterials 2022, 12(19), 3524; https://doi.org/10.3390/nano12193524 - 08 Oct 2022

Cited by 1 | Viewed by 1224

Abstract

For a comprehensive understanding of protein function and dynamics, it is crucial to study their mechanical properties [...] Full article

(This article belongs to the Special Issue Protein Nanomechanics)

Research

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30 pages, 41451 KiB

Open AccessArticle

Exploring the Effect of Mechanical Anisotropy of Protein Structures in the Unfoldase Mechanism of AAA+ Molecular Machines

by Rohith Anand Varikoti, Hewafonsekage Yasan Y. Fonseka, Maria S. Kelly, Alex Javidi, Mangesh Damre, Sarah Mullen, Jimmie L. Nugent IV, Christopher M. Gonzales, George Stan and Ruxandra I. Dima

Nanomaterials 2022, 12(11), 1849; https://doi.org/10.3390/nano12111849 - 28 May 2022

Cited by 5 | Viewed by 2417

Abstract

Essential cellular processes of microtubule disassembly and protein degradation, which span lengths from tens of

μ

m to nm, are mediated by specialized molecular machines with similar hexameric structure and function. Our molecular simulations at atomistic and coarse-grained scales show that both the [...] Read more.

Essential cellular processes of microtubule disassembly and protein degradation, which span lengths from tens of

μ

m to nm, are mediated by specialized molecular machines with similar hexameric structure and function. Our molecular simulations at atomistic and coarse-grained scales show that both the microtubule-severing protein spastin and the caseinolytic protease ClpY, accomplish spectacular unfolding of their diverse substrates, a microtubule lattice and dihydrofolate reductase (DHFR), by taking advantage of mechanical anisotropy in these proteins. Unfolding of wild-type DHFR requires disruption of mechanically strong

β

-sheet interfaces near each terminal, which yields branched pathways associated with unzipping along soft directions and shearing along strong directions. By contrast, unfolding of circular permutant DHFR variants involves single pathways due to softer mechanical interfaces near terminals, but translocation hindrance can arise from mechanical resistance of partially unfolded intermediates stabilized by

β

-sheets. For spastin, optimal severing action initiated by pulling on a tubulin subunit is achieved through specific orientation of the machine versus the substrate (microtubule lattice). Moreover, changes in the strength of the interactions between spastin and a microtubule filament, which can be driven by the tubulin code, lead to drastically different outcomes for the integrity of the hexameric structure of the machine. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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17 pages, 4838 KiB

Open AccessFeature PaperArticle

Nanosurgical Manipulation of Titin and Its M-Complex

by Dominik Sziklai, Judit Sallai, Zsombor Papp, Dalma Kellermayer, Zsolt Mártonfalvi, Ricardo H. Pires and Miklós S. Z. Kellermayer

Nanomaterials 2022, 12(2), 178; https://doi.org/10.3390/nano12020178 - 06 Jan 2022

Cited by 4 | Viewed by 1783

Abstract

Titin is a multifunctional filamentous protein anchored in the M-band, a hexagonally organized supramolecular lattice in the middle of the muscle sarcomere. Functionally, the M-band is a framework that cross-links myosin thick filaments, organizes associated proteins, and maintains sarcomeric symmetry via its structural [...] Read more.

Titin is a multifunctional filamentous protein anchored in the M-band, a hexagonally organized supramolecular lattice in the middle of the muscle sarcomere. Functionally, the M-band is a framework that cross-links myosin thick filaments, organizes associated proteins, and maintains sarcomeric symmetry via its structural and putative mechanical properties. Part of the M-band appears at the C-terminal end of isolated titin molecules in the form of a globular head, named here the “M-complex”, which also serves as the point of head-to-head attachment of titin. We used high-resolution atomic force microscopy and nanosurgical manipulation to investigate the topographical and internal structure and local mechanical properties of the M-complex and its associated titin molecules. We find that the M-complex is a stable structure that corresponds to the transverse unit of the M-band organized around the myosin thick filament. M-complexes may be interlinked into an M-complex array that reflects the local structural and mechanical status of the transversal M-band lattice. Local segments of titin and the M-complex could be nanosurgically manipulated to achieve extension and domain unfolding. Long threads could be pulled out of the M-complex, suggesting that it is a compact supramolecular reservoir of extensible filaments. Nanosurgery evoked an unexpected volume increment in the M-complex, which may be related to its function as a mechanical spacer. The M-complex thus displays both elastic and plastic properties which support the idea that the M-band may be involved in mechanical functions within the muscle sarcomere. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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11 pages, 966 KiB

Open AccessArticle

Force Dependence of Proteins’ Transition State Position and the Bell–Evans Model

by Marc Rico-Pasto, Annamaria Zaltron and Felix Ritort

Nanomaterials 2021, 11(11), 3023; https://doi.org/10.3390/nano11113023 - 11 Nov 2021

Cited by 2 | Viewed by 1941

Abstract

Single-molecule force spectroscopy has opened a new field of research in molecular biophysics and biochemistry. Pulling experiments on individual proteins permit us to monitor conformational transitions with high temporal resolution and measure their free energy landscape. The force–extension curves of single proteins often [...] Read more.

Single-molecule force spectroscopy has opened a new field of research in molecular biophysics and biochemistry. Pulling experiments on individual proteins permit us to monitor conformational transitions with high temporal resolution and measure their free energy landscape. The force–extension curves of single proteins often present large hysteresis, with unfolding forces that are higher than refolding ones. Therefore, the high energy of the transition state (TS) in these molecules precludes kinetic rates measurements in equilibrium hopping experiments. In irreversible pulling experiments, force-dependent kinetic rates measurements show a systematic discrepancy between the sum of the folding and unfolding TS distances derived by the kinetic Bell–Evans model and the full molecular extension predicted by elastic models. Here, we show that this discrepancy originates from the force-induced movement of TS. Specifically, we investigate the highly kinetically stable protein barnase, using pulling experiments and the Bell–Evans model to characterize the position of its kinetic barrier. Experimental results show that while the TS stays at a roughly constant distance relative to the native state, it shifts with force relative to the unfolded state. Interestingly, a conversion of the protein extension into amino acid units shows that the TS position follows the Leffler–Hammond postulate: the higher the force, the lower the number of unzipped amino acids relative to the native state. The results are compared with the quasi-reversible unfolding–folding of a short DNA hairpin. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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11 pages, 2882 KiB

Open AccessArticle

Facile Synthesis of Peptide-Conjugated Gold Nanoclusters with Different Lengths

by Qun Ma, Lichao Liu, Zeyue Yang and Peng Zheng

Nanomaterials 2021, 11(11), 2932; https://doi.org/10.3390/nano11112932 - 02 Nov 2021

Cited by 4 | Viewed by 3025

Abstract

The synthesis of ultra-small gold nanoclusters (Au NCs) with sizes down to 2 nm has received increasing interest due to their unique optical and electronic properties. Like many peptide-coated gold nanospheres synthesized before, modified gold nanoclusters with peptide conjugation are potentially significant in [...] Read more.

The synthesis of ultra-small gold nanoclusters (Au NCs) with sizes down to 2 nm has received increasing interest due to their unique optical and electronic properties. Like many peptide-coated gold nanospheres synthesized before, modified gold nanoclusters with peptide conjugation are potentially significant in biomedical and catalytic fields. Here, we explore whether such small-sized gold nanoclusters can be conjugated with peptides also and characterize them using atomic force microscopy. Using a long and flexible elastin-like polypeptide (ELP)₂₀ as the conjugated peptide, (ELP)₂₀-Au NCs was successfully synthesized via a one-pot synthesis method. The unique optical and electronic properties of gold nanoclusters are still preserved, while a much larger size was obtained as expected due to the peptide conjugation. In addition, a short and rigid peptide (EAAAK)₃ was conjugated to the gold nanoclusters. Their Yong’s modulus was characterized using atomic force microscopy (AFM). Moreover, the coated peptide on the nanoclusters was pulled using AFM-based single molecule-force spectroscopy (SMFS), showing expected properties as one of the first force spectroscopy experiments on peptide-coated nanoclusters. Our results pave the way for further modification of nanoclusters based on the conjugated peptides and show a new method to characterize these materials using AFM-SMFS. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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19 pages, 4857 KiB

Open AccessEditor’s ChoiceArticle

Interpretation of Single-Molecule Force Experiments on Proteins Using Normal Mode Analysis

by Jacob Bauer and Gabriel Žoldák

Nanomaterials 2021, 11(11), 2795; https://doi.org/10.3390/nano11112795 - 22 Oct 2021

Cited by 1 | Viewed by 1521

Abstract

Single-molecule force spectroscopy experiments allow protein folding and unfolding to be explored using mechanical force. Probably the most informative technique for interpreting the results of these experiments at the structural level makes use of steered molecular dynamics (MD) simulations, which can explicitly model [...] Read more.

Single-molecule force spectroscopy experiments allow protein folding and unfolding to be explored using mechanical force. Probably the most informative technique for interpreting the results of these experiments at the structural level makes use of steered molecular dynamics (MD) simulations, which can explicitly model the protein under load. Unfortunately, this technique is computationally expensive for many of the most interesting biological molecules. Here, we find that normal mode analysis (NMA), a significantly cheaper technique from a computational perspective, allows at least some of the insights provided by MD simulation to be gathered. We apply this technique to three non-homologous proteins that were previously studied by force spectroscopy: T4 lysozyme (T4L), Hsp70 and the glucocorticoid receptor domain (GCR). The NMA results for T4L and Hsp70 are compared with steered MD simulations conducted previously, and we find that we can recover the main results. For the GCR, which did not undergo MD simulation, our approach identifies substructures that correlate with experimentally identified unfolding intermediates. Overall, we find that NMA can make a valuable addition to the analysis toolkit for the structural analysis of single-molecule force experiments on proteins. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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19 pages, 4704 KiB

Open AccessArticle

Classifying Residues in Mechanically Stable and Unstable Substructures Based on a Protein Sequence: The Case Study of the DnaK Hsp70 Chaperone

by Michal Gala and Gabriel Žoldák

Nanomaterials 2021, 11(9), 2198; https://doi.org/10.3390/nano11092198 - 26 Aug 2021

Cited by 3 | Viewed by 2596

Abstract

Artificial proteins can be constructed from stable substructures, whose stability is encoded in their protein sequence. Identifying stable protein substructures experimentally is the only available option at the moment because no suitable method exists to extract this information from a protein sequence. In [...] Read more.

Artificial proteins can be constructed from stable substructures, whose stability is encoded in their protein sequence. Identifying stable protein substructures experimentally is the only available option at the moment because no suitable method exists to extract this information from a protein sequence. In previous research, we examined the mechanics of E. coli Hsp70 and found four mechanically stable (S class) and three unstable substructures (U class). Of the total 603 residues in the folded domains of Hsp70, 234 residues belong to one of four mechanically stable substructures, and 369 residues belong to one of three unstable substructures. Here our goal is to develop a machine learning model to categorize Hsp70 residues using sequence information. We applied three supervised methods: logistic regression (LR), random forest, and support vector machine. The LR method showed the highest accuracy, 0.925, to predict the correct class of a particular residue only when context-dependent physico-chemical features were included. The cross-validation of the LR model yielded a prediction accuracy of 0.879 and revealed that most of the misclassified residues lie at the borders between substructures. We foresee machine learning models being used to identify stable substructures as candidates for building blocks to engineer new proteins. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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Review

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10 pages, 2045 KiB

Open AccessReview

Bioconjugation Strategies for Connecting Proteins to DNA-Linkers for Single-Molecule Force-Based Experiments

by Lyan M. van der Sleen and Katarzyna M. Tych

Nanomaterials 2021, 11(9), 2424; https://doi.org/10.3390/nano11092424 - 17 Sep 2021

Cited by 10 | Viewed by 4209

Abstract

The mechanical properties of proteins can be studied with single molecule force spectroscopy (SMFS) using optical tweezers, atomic force microscopy and magnetic tweezers. It is common to utilize a flexible linker between the protein and trapped probe to exclude short-range interactions in SMFS [...] Read more.

The mechanical properties of proteins can be studied with single molecule force spectroscopy (SMFS) using optical tweezers, atomic force microscopy and magnetic tweezers. It is common to utilize a flexible linker between the protein and trapped probe to exclude short-range interactions in SMFS experiments. One of the most prevalent linkers is DNA due to its well-defined properties, although attachment strategies between the DNA linker and protein or probe may vary. We will therefore provide a general overview of the currently existing non-covalent and covalent bioconjugation strategies to site-specifically conjugate DNA-linkers to the protein of interest. In the search for a standardized conjugation strategy, considerations include their mechanical properties in the context of SMFS, feasibility of site-directed labeling, labeling efficiency, and costs. Full article

(This article belongs to the Special Issue Protein Nanomechanics)

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