Improving the Performance of a Quadrupole Time-of-Flight Instrument for Macromolecular Mass Spectrometry

Heuvel, Robert H. H. van den; Duijn, Esther van; Mazon, Hortense; Synowsky, Silvia; Lorenzen, Kristina; Versluis, Cees; Brouns, Stan J. J.; Langridge, Dave; Oost, John van der; Hoyes, John; Heck, Albert J. R.

doi:10.1021/ac061039a

Cited by 244 publications

(274 citation statements)

References 42 publications

Supporting

Mentioning

261

Contrasting

Unclassified

Order By: Relevance

“…Needles were made from borosilicate glass capillaries (Kwik-Fil, World Precision Instruments, Sarasota, FL) on a P97 puller (Sutter Instruments, Novato, CA), coated with a thin gold layer by using an Edwards Scancoat six pirani 501 sputter coater (Edwards Laboratories, Milpitas, USA). The instruments were adjusted for optimal performance in high mass detection (32).…”

Section: Mass Spectrometry Of Cascade and Csymentioning

confidence: 99%

“…For tandem mass spectrometric analysis xenon was used as the collision gas at a pressure of 1.5 10 Ϫ2 mbar on the modified Q-ToF1 (32). The collision voltage varied between 10 -200V to monitor the sequential loss of several Cse/Cas or Csy proteins from the intact Cascade or Csy machinery respectively, or its subcomplexes.…”

Section: Mass Spectrometry Of Cascade and Csymentioning

confidence: 99%

See 1 more Smart Citation

Native Tandem and Ion Mobility Mass Spectrometry Highlight Structural and Modular Similarities in Clustered-Regularly-Interspaced Shot-Palindromic-Repeats (CRISPR)-associated Protein Complexes From Escherichia coli and Pseudomonas aeruginosa

Duijn

Barbu

Barendregt

et al. 2012

Molecular & Cellular Proteomics

Self Cite

View full text Add to dashboard Cite

The CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated genes) immune system of bacteria and archaea provides acquired resistance against viruses and plasmids, by a strategy analogous to RNA-interference. Key components of the defense system are ribonucleoprotein complexes, the composition of which appears highly variable in different CRISPR/Cas subtypes. Previous studies combined mass spectrometry, electron microscopy, and small angle x-ray scattering to demonstrate that the E. coli Cascade complex (405 kDa) and the P. aeruginosa Csy-complex (350 kDa) are similar in that they share a central spiral-shaped hexameric structure, flanked by associating proteins and one CRISPR RNA. Recently, a cryo-electron microscopy structure of Cascade revealed that the CRISPR RNA molecule resides in a groove of the hexameric backbone. For both complexes we here describe the use of native mass spectrometry in combination with ion mobility mass spectrometry to assign a stable core surrounded by more loosely associated modules. Via computational modeling subcomplex structures were proposed that relate to the experimental IMMS data. Despite the absence of obvious sequence homology between several subunits, detailed analysis of sub-complexes strongly suggests analogy between subunits of the two complexes. Probing the specific association of E. coli Cascade/crRNA to its complementary DNA target reveals a conformational change. All together these findings provide relevant new information about the potential assembly process of the two CRISPR-associated complexes. Molecular & Cellular

show abstract

Section: Mass Spectrometry Of Cascade and Csymentioning

confidence: 99%

Section: Mass Spectrometry Of Cascade and Csymentioning

confidence: 99%

Native Tandem and Ion Mobility Mass Spectrometry Highlight Structural and Modular Similarities in Clustered-Regularly-Interspaced Shot-Palindromic-Repeats (CRISPR)-associated Protein Complexes From Escherichia coli and Pseudomonas aeruginosa

Duijn

Barbu

Barendregt

et al. 2012

Molecular & Cellular Proteomics

Self Cite

View full text Add to dashboard Cite

show abstract

“…Heck and coworkers [44] reported interesting dissociation behaviour for the dissociation of tetrameric complexes of 2-keto-3-deoxyarabinonate dehydratase. These complexes dissociate almost exclusively into two dimeric fragment ions rather than the usual loss of a single monomer.…”

Section: Application Of the Coulomb Repulsion Modelmentioning

confidence: 99%

Theoretical investigations of the dissociation of charged protein complexes in the gas phase

Wanasundara

Thachuk

2007

J. Am. Soc. Mass Spectrom.

View full text Add to dashboard Cite

A series of calculations, varying from simple electrostatic to more detailed semi-empirical based molecular dynamics ones, were carried out on charged gas phase ions of the cytochrome c dimer. The energetics of differing charge states, charge partitionings, and charge configurations were examined in both the low and high charge regimes. As well, preliminary free energy calculations of dissociation barriers are presented. It is shown that one must always consider distributions of charge configurations, once protein relaxation effects are taken into account, and that no single configuration dominates. All these results also indicate that in the high charge limit, the dissociation of protein complex ions is governed by electrostatic repulsion from the net charges, the consequences of which are enumerated and discussed. There are two main trends deriving from this, namely that charges will move so as to approximately maintain constant surface charge density, and that the lowest barrier to dissociation is the one that produces fragment ions with equal charges. In particular, it is shown that the charge-to-mass ratio of a fragment ion is not the key physical parameter in predicting dissociation products. In fact, from the perspective of the division of total charge, many dissociation pathways reported to be "asymmetric" in the literature should be more properly labelled as "symmetric" or "near-symmetric". The Coulomb repulsion model assumes that the timescale for charge transfer is faster than that for protein structural changes, which in turn is faster than that for complex dissociation. One challenge in using mass spectrometry for the general analysis of protein complexes is understanding the dissociation mechanism of these complexes in the gas phase. Many groups [5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] have reported asymmetric dissociation behaviour for large multimeric proteins. A small subunit, typically a protein monomer, is ejected from the complex during dissociation, with the monomer carrying away a disproportionate amount of charge for its relative mass. Understanding the cause of this phenomenon will help to improve our understanding of the structural properties of noncovalent complexes.It is important to investigate how these dissociations occur and the factors that influence them. Using Fourier-transform mass spectrometry (FTMS), Jurchen and Williams [6] have conducted a number of studies in order to understand the origin of these charge distributions. In these experiments, isolated charge states of cytochrome c dimer were dissociated by sustained off-resonance irradiation collisionally activated dissociation (SORI-CAD). According to Jurchen and Williams [6], the asymmetric charge distribution depends upon charge state, dissociation energy, and conformational flexibility. These studies showed that higher charge states lead to symmetric charge products while lower charge states lead to an asymmetric charge dissociation pathway. Further, their results with different excitation energies show ...

show abstract

“…From adjacent signals in a charge state distribution, the mass can be obtained; in practice, all ion signals are used for a more accurate determination. Recent developments in Q-TOF mass spectrometers (18,25) have enabled the analyses of species with masses over 1 million Da (16,23,26,27). The connector protein was analyzed previously using native mass spectrometry (16) and found to be dodecameric.…”

Section: Characterization Of Connector-scaffolding Complexes-mentioning

confidence: 99%

A Docking Model Based on Mass Spectrometric and Biochemical Data Describes Phage Packaging Motor Incorporation

Uetrecht

Kang

et al. 2010

Molecular & Cellular Proteomics

View full text Add to dashboard Cite

The molecular mechanism of scaffolding protein-mediated incorporation of one and only one DNA packaging motor/connector dodecamer at a unique vertex during lambdoid phage assembly has remained elusive because of the lack of structural information on how the connector and scaffolding proteins interact. We assembled and characterized a 29 connector-scaffolding complex, which can be incorporated into procapsids during in vitro assembly. Native mass spectrometry revealed that the connector binds at most 12 scaffolding molecules, likely organized as six dimers. A data-driven docking model, using input from chemical cross-linking and mutagenesis data, suggested an interaction between the scaffolding protein and the exterior of the wide domain of the connector dodecamer. The connector binding region of the scaffolding protein lies upstream of the capsid binding region located at the C terminus. This arrangement allows the C terminus of scaffolding protein within the complex to both recruit capsid subunits and mediate the incorporation of the single connector vertex. Molecular & Cellular Proteomics 9:1764 -1773, 2010.The DNA packaging motor of double-stranded DNA bacteriophages translocates genomic DNA into a preformed procapsid to near crystalline density and is the strongest motor characterized to date. The packaging motor of the Bacillus subtilis phage 29 can work against 57 piconewtons of internal force and translocate 2 bp of DNA per ATP hydrolyzed at a maximum velocity of 103 bp/s (1, 2). The motor complex is assembled on a dodecamer of the connector protein, which replaces a pentameric vertex in the procapsid and serves both as a portal for DNA passage and the docking site for the other packaging components (3).To successfully package a full-length genome, incorporation of one and only one connector vertex is essential (4). In vivo, nearly every assembled procapsid has one and only one connector vertex and is able to package DNA and mature into an infectious phage (5). This narrow distribution in which 95% of particles have a single connector vertex cannot be explained by random statistical incorporation. The control mechanism is coupled to the procapsid assembly process. Procapsid assembly requires the copolymerization of hundreds of copies each of the capsid and scaffolding proteins as well as a dodecamer of the portal or connector protein. The scaffolding protein acts to both activate the coat protein for assembly and ensure proper form determination. In the absence of scaffolding protein, uncontrolled polymerization results in the assembly of aberrant structures. In a properly assembled procapsid, the portal protein is located at one vertex, whereas scaffolding protein occupies the bulk of the interior space and is subsequently removed during DNA packaging by either proteolysis or simple release. Mutational studies have indicated that scaffolding protein is involved either directly or indirectly in the incorporation of the connector vertex during procapsid assembly in a variety of phages (6 -8).In 29, the con...

show abstract

Improving the Performance of a Quadrupole Time-of-Flight Instrument for Macromolecular Mass Spectrometry

Cited by 244 publications

References 42 publications

Native Tandem and Ion Mobility Mass Spectrometry Highlight Structural and Modular Similarities in Clustered-Regularly-Interspaced Shot-Palindromic-Repeats (CRISPR)-associated Protein Complexes From Escherichia coli and Pseudomonas aeruginosa

Native Tandem and Ion Mobility Mass Spectrometry Highlight Structural and Modular Similarities in Clustered-Regularly-Interspaced Shot-Palindromic-Repeats (CRISPR)-associated Protein Complexes From Escherichia coli and Pseudomonas aeruginosa

Theoretical investigations of the dissociation of charged protein complexes in the gas phase

A Docking Model Based on Mass Spectrometric and Biochemical Data Describes Phage Packaging Motor Incorporation

Contact Info

Product

Resources

About