Probing the Structure of the Caulobacter crescentus Ribosome with Chemical Labeling and Mass Spectrometry

Beardsley, Richard L.; Running, William E.; Reilly, James P.

doi:10.1021/pr060170w

Cited by 30 publications

(58 citation statements)

References 55 publications

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“…For example, the N-terminus of the poliovirus capsid protein VP1 that is entirely internal in native virion becomes externalized upon cell attachment, and the exposed N-terminus of VP1 was shown to be responsible for liposome binding [10, 61]. These conformational changes can be predicted from our simulations and used to guide experimental studies, e.g., chemical labeling[62]. If the protein terminal domains are protruding/retracting in a fluctuating manner, then the degree of labeling will increase when such fluctuations are large.…”

Section: Discussionmentioning

confidence: 99%

All-Atom Multiscale Simulation of Cowpea Chlorotic Mottle Virus Capsid Swelling

2010

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An all-atom multiscale computational modeling approach, Molecular Dynamics/Order Parameter eXtrapolation (MD/OPX), has recently been developed for simulating large bionanosystems. It accelerates MD simulations and addresses rapid atomistic fluctuations and slowly-varying nanoscale dynamics of bionanosystems simultaneously. With modules added to account for water molecules and ions, MD/OPX is applied to simulate the swelling of cowpea chlorotic mottle virus (CCMV) capsid solvated in a host medium in this study. Simulation results show that the Nterminal arms of capsid proteins undergo large deviations from the initial configurations with their length extended quickly during the early stage of capsid swelling. The capsid swelling is a symmetry-breaking process involving local initiation and front propagation. The capsid swelling rate is ~0.25 nm/ns (npn) during early stage of the simulation and propagation of the structural transition across the capsid is roughly 0.6npn. The system conditions that affect swelling of the capsid are analyzed. Prospects for creating a phase diagram for CCMV capsid swelling and using predictions to guide experiments are discussed.

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Section: Discussionmentioning

confidence: 99%

All-Atom Multiscale Simulation of Cowpea Chlorotic Mottle Virus Capsid Swelling

2010

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“…Unlike most other lysine-modification reagents, their studies revealed that these labels can also modify cysteine residues. Using the same reagents, the structure of the Caulobacter crescentus ribosome was characterized (Beardsley, Running, & Reilly, 2006). The ribosomal proteins were amidinated before and after ribosome disassembly.…”

Section: Amino Acid-specific Labels and Examples Of Their Applicmentioning

confidence: 99%

Probing protein structure by amino acid‐specific covalent labeling and mass spectrometry

Mendoza

Vachet

2008

Mass Spectrometry Reviews

327

445

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For many years, amino acid-specific covalent labeling has been a valuable tool to study protein structure and protein interactions, especially for systems that are difficult to study by other means. These covalent labeling methods typically map protein structure and interactions by measuring the differential reactivity of amino acid side chains. The reactivity of amino acids in proteins generally depends on the accessibility of the side chain to the reagent, the inherent reactivity of the label and the reactivity of the amino acid side chain. Peptide mass mapping with ESI- or MALDI-MS and peptide sequencing with tandem MS are typically employed to identify modification sites to provide site-specific structural information. In this review, we describe the reagents that are most commonly used in these residue-specific modification reactions, details about the proper use of these covalent labeling reagents, and information about the specific biochemical problems that have been addressed with covalent labeling strategies.

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“…Future experiments capable of investigating the high resolution structure (Beardsley et al, 2006; Namy et al, 2006; Selmer et al, 2006) and/or real-time conformational dynamics (Blanchard et al, 2004; Cornish et al, 2008; Lee et al, 2007; Munro et al, 2007) of single ribosome-mRNA-tRNA complexes paused over a frameshifting signal hold considerable promise for enhancing our mechanistic understanding of how programmed ribosomal recoding events function (for a recent review on single molecule methods as applied to translation, see Munro et al, 2008). For example, a recent study reports the use of optical trapping methods to observe single ribosomes translocating through an RNA stem-loop structure via continuous measurement of the end-to-end length of the mRNA as it is unwound by the ribosome (Wen et al, 2008).…”

Section: Future Prospectivementioning

confidence: 99%

Frameshifting RNA pseudoknots: Structure and mechanism

2009

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Programmed ribosomal frameshifting (PRF) is one of multiple translational recoding processes that fundamentally alters triplet decoding of the messenger RNA by the elongating ribosome. The ability of the ribosome to change translational reading frames in the −1 direction (−1 PRF) is employed by many positive strand RNA viruses, including economically important plant viruses and many human pathogens such as retroviruses, e.g., HIV-1, and coronaviruses, e.g., the causative agent of severe acute respiratory syndrome (SARS), in order to properly express their genomes. −1 PRF is programmed by a bipartite signal embedded in the mRNA and includes a heptanucleotide "slip site" over which the paused ribosome "backs up" by one nucleotide, and a downstream stimulatory element, either an RNA pseudoknot or a very stable RNA stem-loop. These two elements are separated by 6-8 nucleotides, a distance that places the 5′ edge of the downstream stimulatory element in direct contact with the mRNA entry channel of the 30S ribosomal subunit. The precise mechanism by which the downstream RNA stimulates −1 PRF by the translocating ribosome remains unclear. This review summarizes the recent structural and biophysical studies of RNA pseudoknots and places this work in the context of our evolving mechanistic understanding of translation elongation. Support for the hypothesis that the downstream stimulatory element provides a kinetic barrier to the ribosome mediated unfolding is discussed.

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Probing the Structure of the Caulobacter crescentus Ribosome with Chemical Labeling and Mass Spectrometry

Cited by 30 publications

References 55 publications

All-Atom Multiscale Simulation of Cowpea Chlorotic Mottle Virus Capsid Swelling

All-Atom Multiscale Simulation of Cowpea Chlorotic Mottle Virus Capsid Swelling

Probing protein structure by amino acid‐specific covalent labeling and mass spectrometry

Frameshifting RNA pseudoknots: Structure and mechanism

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