liu.seSearch for publications in DiVA
Change search
Link to record
Permanent link

Direct link
BETA
Publications (10 of 10) Show all publications
Niklasson, M., Otten, R., Ahlner, A., Andrésen, C., Schlagnitweit, J., Petzold, K. & Lundström, P. (2017). Comprehensive analysis of NMR data using advanced line shape fitting.. Journal of Biomolecular NMR, 69(2), 93-99
Open this publication in new window or tab >>Comprehensive analysis of NMR data using advanced line shape fitting.
Show others...
2017 (English)In: Journal of Biomolecular NMR, ISSN 0925-2738, E-ISSN 1573-5001, Vol. 69, no 2, p. 93-99Article in journal (Refereed) Published
Abstract [en]

NMR spectroscopy is uniquely suited for atomic resolution studies of biomolecules such as proteins, nucleic acids and metabolites, since detailed information on structure and dynamics are encoded in positions and line shapes of peaks in NMR spectra. Unfortunately, accurate determination of these parameters is often complicated and time consuming, in part due to the need for different software at the various analysis steps and for validating the results. Here, we present an integrated, cross-platform and open-source software that is significantly more versatile than the typical line shape fitting application. The software is a completely redesigned version of PINT ( https://pint-nmr.github.io/PINT/ ). It features a graphical user interface and includes functionality for peak picking, editing of peak lists and line shape fitting. In addition, the obtained peak intensities can be used directly to extract, for instance, relaxation rates, heteronuclear NOE values and exchange parameters. In contrast to most available software the entire process from spectral visualization to preparation of publication-ready figures is done solely using PINT and often within minutes, thereby, increasing productivity for users of all experience levels. Unique to the software are also the outstanding tools for evaluating the quality of the fitting results and extensive, but easy-to-use, customization of the fitting protocol and graphical output. In this communication, we describe the features of the new version of PINT and benchmark its performance.

Place, publisher, year, edition, pages
Springer, 2017
Keywords
Dynamics, Line shape fitting, Peak integration, Relaxation, Spectral analysis
National Category
Biochemistry and Molecular Biology Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-142786 (URN)10.1007/s10858-017-0141-6 (DOI)000414206400004 ()29043470 (PubMedID)2-s2.0-85031497711 (Scopus ID)
Note

Funding agencies: Swedish Research Council [2012-5136]

Available from: 2017-11-03 Created: 2017-11-03 Last updated: 2017-12-04Bibliographically approved
Niklasson, M., Ahlner, A., Andrésen, C., Marsh, J. A. & Lundström, P. (2015). Fast and Accurate Resonance Assignment of Small-to-Large Proteins by Combining Automated and Manual Approaches. PloS Computational Biology, 11(1), e1004022
Open this publication in new window or tab >>Fast and Accurate Resonance Assignment of Small-to-Large Proteins by Combining Automated and Manual Approaches
Show others...
2015 (English)In: PloS Computational Biology, ISSN 1553-734X, E-ISSN 1553-7358, Vol. 11, no 1, p. e1004022-Article in journal (Refereed) Published
Abstract [en]

The process of resonance assignment is fundamental to most NMR studies of protein structure and dynamics. Unfortunately, the manual assignment of residues is tedious and time-consuming, and can represent a significant bottleneck for further characterization. Furthermore, while automated approaches have been developed, they are often limited in their accuracy, particularly for larger proteins. Here, we address this by introducing the software COMPASS, which, by combining automated resonance assignment with manual intervention, is able to achieve accuracy approaching that from manual assignments at greatly accelerated speeds. Moreover, by including the option to compensate for isotope shift effects in deuterated proteins, COMPASS is far more accurate for larger proteins than existing automated methods. COMPASS is an open-source project licensed under GNU General Public License and is available for download from http://www.liu.se/forskning/foass/tidigare-foass/patrik-lundstrom/software?l=en. Source code and binaries for Linux, Mac OS X and Microsoft Windows are available.

Place, publisher, year, edition, pages
Public Library of Science, 2015
National Category
Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-115010 (URN)10.1371/journal.pcbi.1004022 (DOI)000349309400013 ()25569628 (PubMedID)
Note

Funding Agencies|Swedish Research Council [Dnr. 2012-5136]

Available from: 2015-03-09 Created: 2015-03-06 Last updated: 2017-12-04
Ahlner, A., Andresen, C., Khan, S. N., Kay, L. E. & Lundström, P. (2015). Fractional enrichment of proteins using [2-13C]-glycerol as the carbon source facilitates measurement of excited state 13Cα chemical shifts with improved sensitivity. Journal of Biomolecular NMR, 62(3), 341-351
Open this publication in new window or tab >>Fractional enrichment of proteins using [2-13C]-glycerol as the carbon source facilitates measurement of excited state 13Cα chemical shifts with improved sensitivity
Show others...
2015 (English)In: Journal of Biomolecular NMR, ISSN 0925-2738, E-ISSN 1573-5001, Vol. 62, no 3, p. 341-351Article in journal (Refereed) Published
Abstract [en]

A selective isotope labeling scheme based on the utilization of [2-13C]-glycerol as the carbon source during protein overexpression has been evaluated for the measurement of excited state 13Cα chemical shifts using Carr–Purcell–Meiboom–Gill (CPMG) relaxation dispersion (RD) experiments. As expected, the fractional incorporation of label at the Cα positions is increased two-fold relative to labeling schemes based on [2-13C]-glucose, effectively doubling the sensitivity of NMR experiments. Applications to a binding reaction involving an SH3 domain from the protein Abp1p and a peptide from the protein Ark1p establish that accurate excited state 13Cα chemical shifts can be obtained from RD experiments, with errors on the order of 0.06 ppm for exchange rates ranging from 100 to 1000 s−1, despite the small fraction of 13Cα–13Cβ spin-pairs that are present for many residue types. The labeling approach described here should thus be attractive for studies of exchanging systems using 13Cα spin probes.

Place, publisher, year, edition, pages
Springer Netherlands, 2015
Keywords
CPMG, 13Cα labeling, [2-13C]-Glycerol, Excited states
National Category
Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-117073 (URN)10.1007/s10858-015-9948-1 (DOI)000357489200010 ()
Note

At the time for thesis presentation publication was in status: Manuscript

At the time for thesis presentation name of publication was: Fractional enrichment using [2-13C]-glycerol as the carbon source facilitates measurements of excited state 13Cα chemical shifts with improved sensitivity

Available from: 2015-04-15 Created: 2015-04-15 Last updated: 2017-12-04Bibliographically approved
Ahlner, A. (2015). Improved Methods for Characterization of Protein Dynamics by NMR spectroscopy and Studies of the EphB2 Kinase Domain. (Doctoral dissertation). Linköping: Linköping University Electronic Press
Open this publication in new window or tab >>Improved Methods for Characterization of Protein Dynamics by NMR spectroscopy and Studies of the EphB2 Kinase Domain
2015 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Proteins are essential for all known forms of life and in many lethal diseases protein failure is the cause of the disease. To understand proteins and the processes they are involved in, it is valuable to know their structures as well as their dynamics and interactions. The structures may not be directly inspected because proteins are too small to be visible in a light microscope, which is why indirect methods such as nuclear magnetic resonance (NMR) spectroscopy have to be utilized. This method provides atomic information about the protein and, in contrast to other methods with similar resolution, the measurements are performed in solution resulting in more physiological conditions, enabling analysis of dynamics. Important dynamical processes are the ones on the millisecond timeframe, which may contribute to interactions of proteins and their catalysis of chemical reactions, both of significant value for the function of the proteins.

To better understand proteins, not only do we need to study them, but also develop the methods we are using. This thesis presents four papers about improved NMR techniques as well as a fifth where the kinase domain of ephrinB receptor 2 (EphB2) has been studied regarding the importance of millisecond dynamics and interactions for the activation process. The first paper presents the software COMPASS, which combines statistics and the calculation power of a computer with the flexibility and experience of the user to facilitate and speed up the process of assigning NMR signals to the atoms in the protein. The computer program PINT has been developed for easier and faster evaluation of NMR experiments, such as those that evaluate protein dynamics. It is especially helpful for NMR signals that are difficult to distinguish, so called overlapped peaks, and the soft- ware also converts the detected signals to the indirectly measured physical quantities, such as relaxation rate constants, principal for dynamics. Next are two new versions of the Carr-Purcell-Maiboom-Gill (CPMG) dispersion pulse sequences, designed to measure millisecond dynamics in a way so that the signals are more separated than in standard experiments, to reduce problems with overlaps. To speed up the collection time of the data set, a subset is collected and the entire data set is then reconstructed, by multi-dimensional decomposition co-processing. Described in the thesis is also a way to produce suitably labeled proteins, to detect millisecond dynamics at Cα positions in proteins, using the CPMG dispersion relaxation experiment at lower protein concentrations. Lastly, the kinase domain of EphB2 is shown to be more dynamic on the millisecond time scale as well as more prone to interact with itself in the active form than in the inactive one. This is important for the receptor function of the protein, when and how it mediates signals.

To conclude, this work has extended the possibilities to study protein dynamics by NMR spectroscopy and contributed to increased understanding of the activation process of EphB2 and its signaling mechanism. 

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2015. p. 58
Series
Linköping Studies in Science and Technology. Dissertations, ISSN 0345-7524 ; 1649
National Category
Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-117076 (URN)10.3384/diss.diva-117076 (DOI)978-91-7519-103-4 (ISBN)
Public defence
2015-05-22, Planck, Fysikhuset, Campus Valla, Linköping, 09:15 (English)
Opponent
Supervisors
Available from: 2015-04-15 Created: 2015-04-15 Last updated: 2019-11-15Bibliographically approved
Ahlner, A., Carlsson, M., Jonsson, B.-H. & Lundström, P. (2013). PINT: a software for integration of peak volumes and extraction of relaxation rates. Journal of Biomolecular NMR, 56(3), 191-202
Open this publication in new window or tab >>PINT: a software for integration of peak volumes and extraction of relaxation rates
2013 (English)In: Journal of Biomolecular NMR, ISSN 0925-2738, E-ISSN 1573-5001, Vol. 56, no 3, p. 191-202Article in journal (Refereed) Published
Abstract [en]

We present the software Peak INTegration (PINT), designed to perform integration of peaks in NMR spectra. The program is very simple to run, yet powerful enough to handle complicated spectra. Peaks are integrated by fitting predefined line shapes to experimental data and the fitting can be customized to deal with, for instance, heavily overlapped peaks. The results can be inspected visually, which facilitates systematic optimization of the line shape fitting. Finally, integrated peak volumes can be used to extract parameters such as relaxation rates and information about low populated states. The utility of PINT is demonstrated by applications to the 59 residue SH3 domain of the yeast protein Abp1p and the 289 residue kinase domain of murine EphB2.

Place, publisher, year, edition, pages
Springer Verlag (Germany), 2013
Keywords
Peak integration, Overlapped peaks, Relaxation rates, Protein dynamics
National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:liu:diva-95951 (URN)10.1007/s10858-013-9737-7 (DOI)000321544600001 ()
Note

Funding Agencies|Swedish Research Council||

Available from: 2013-08-19 Created: 2013-08-12 Last updated: 2018-01-11Bibliographically approved
Weininger, U., Blissing, A. T., Hennig, J., Ahlner, A., Liu, Z., Vogel, H. J., . . . Lundström, P. (2013). Protein conformational exchange measured by H-1 R-1 rho relaxation dispersion of methyl groups. Journal of Biomolecular NMR, 57(1), 47-55
Open this publication in new window or tab >>Protein conformational exchange measured by H-1 R-1 rho relaxation dispersion of methyl groups
Show others...
2013 (English)In: Journal of Biomolecular NMR, ISSN 0925-2738, E-ISSN 1573-5001, Vol. 57, no 1, p. 47-55Article in journal (Refereed) Published
Abstract [en]

Activated dynamics plays a central role in protein function, where transitions between distinct conformations often underlie the switching between active and inactive states. The characteristic time scales of these transitions typically fall in the microsecond to millisecond range, which is amenable to investigations by NMR relaxation dispersion experiments. Processes at the faster end of this range are more challenging to study, because higher RF field strengths are required to achieve refocusing of the exchanging magnetization. Here we describe a rotating-frame relaxation dispersion experiment for H-1 spins in methyl (CHD2)-C-13 groups, which improves the characterization of fast exchange processes. The influence of H-1-H-1 rotating-frame nuclear Overhauser effects (ROE) is shown to be negligible, based on a comparison of R (1 rho) relaxation data acquired with tilt angles of 90A degrees and 35A degrees, in which the ROE is maximal and minimal, respectively, and on samples containing different H-1 densities surrounding the monitored methyl groups. The method was applied to ubiquitin and the apo form of calmodulin. We find that ubiquitin does not exhibit any H-1 relaxation dispersion of its methyl groups at 10 or 25 A degrees C. By contrast, calmodulin shows significant conformational exchange of the methionine methyl groups in its C-terminal domain, as previously demonstrated by H-1 and C-13 CPMG experiments. The present R (1 rho) experiment extends the relaxation dispersion profile towards higher refocusing frequencies, which improves the definition of the exchange correlation time, compared to previous results.

Place, publisher, year, edition, pages
Springer Verlag (Germany), 2013
Keywords
Relaxation dispersion, Conformational exchange, Rotating-frame relaxation
National Category
Engineering and Technology
Identifiers
urn:nbn:se:liu:diva-97653 (URN)10.1007/s10858-013-9764-4 (DOI)000323673800006 ()
Note

Funding Agencies|Swedish Research Council|621-2010-4912822-2005-2915621-2012-5136|Goran Gustafsson Foundation for Research in Natural Sciences and Medicine||Knut and Alice Wallenberg Foundation||EMBO long-term fellowship||Forum Scientium Award||Alberta Innovates Health Solutions||

Available from: 2013-09-19 Created: 2013-09-19 Last updated: 2017-12-06
Lundström, P., Ahlner, A. & Blissing, A. T. (2012). Isotope labeling methods for large systems. In: Hanudatta S. Atreya (Ed.), Isotope labeling in Biomolecular NMR: (pp. 3-15). Springer
Open this publication in new window or tab >>Isotope labeling methods for large systems
2012 (English)In: Isotope labeling in Biomolecular NMR / [ed] Hanudatta S. Atreya, Springer, 2012, p. 3-15Chapter in book (Refereed)
Abstract [en]

NMR spectroscopy has undergone a revolution in recent years with the advent of several new methods overcoming the problems of sensitivity and resolution. Recent developments in biotechnology have made it easier and economical to introduce 13C, 15N and 2H into proteins and nucleic acids. At the same time, there has been an explosion in the number of NMR experiments that utilize such isotope labeled samples. Thus, a combination of isotopic labeling and multidimensional, multinuclear NMR has opened up new avenues for structural studies of proteins, nucleic acids and their complexes. This book wil

Place, publisher, year, edition, pages
Springer, 2012
Series
Advances in Experimental Medicine and Biology, ISSN 0065-2598 ; 992
Keywords
Nuclear magnetic resonance spectroscopy, Radiolabeling, Biomolecules- Analysis, Isotope Labeling -methods, Nuclear Magnetic Resonance, Biomolecular
National Category
Natural Sciences
Identifiers
urn:nbn:se:liu:diva-85053 (URN)10.1007/978-94-007-4954-2_1 (DOI)978-94-007-4953-5 (ISBN)978-94-007-4954-2 (ISBN)
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2015-04-15Bibliographically approved
Lundström, P., Ahlner, A. & Blissing, A. T. (2012). Isotope labeling methods for relaxation measurements. In: Hanudatta S. Atreya (Ed.), Isotope labeling in Biomolecular NMR: (pp. 63-82). Springer
Open this publication in new window or tab >>Isotope labeling methods for relaxation measurements
2012 (English)In: Isotope labeling in Biomolecular NMR / [ed] Hanudatta S. Atreya, Springer, 2012, p. 63-82Chapter in book (Refereed)
Abstract [en]

NMR spectroscopy has undergone a revolution in recent years with the advent of several new methods overcoming the problems of sensitivity and resolution. Recent developments in biotechnology have made it easier and economical to introduce 13C, 15N and 2H into proteins and nucleic acids. At the same time, there has been an explosion in the number of NMR experiments that utilize such isotope labeled samples. Thus, a combination of isotopic labeling and multidimensional, multinuclear NMR has opened up new avenues for structural studies of proteins, nucleic acids and their complexes. This book wil

Place, publisher, year, edition, pages
Springer, 2012
Series
Advances in Experimental Medicine and Biology, ISSN 0065-2598 ; 992
Keywords
Nuclear magnetic resonance spectroscopy, Radiolabeling, Biomolecules-Analysis, Isotope Labeling-methods, Nuclear Magnetic Resonance, Biomolecular
National Category
Natural Sciences
Identifiers
urn:nbn:se:liu:diva-85052 (URN)10.1007/978-94-007-4954-2_4 (DOI)978-94-007-4953-5 (ISBN)978-94-007-4954-2 (ISBN)
Available from: 2012-10-31 Created: 2012-10-31 Last updated: 2015-04-15Bibliographically approved
Ahlner, A., Khan, S. N., Forman-Kay, J. D., Sicheri, F. & Lundström, P.Conformational Dynamics and Multimerization of Active Forms of the EphrinB Receptor 2 Kinase Domain.
Open this publication in new window or tab >>Conformational Dynamics and Multimerization of Active Forms of the EphrinB Receptor 2 Kinase Domain
Show others...
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Active and autoinhibited forms of the ephrinB receptor 2 (EphB2) kinase domain have been studied using NMR spectroscopy. The project was initiated because of the finding that the crystal structures of active forms of the kinase domain and previous NMR studies suggested that a change in inter-lobe flexibility and the sampling of catalytically competent excited states conformations are responsible for activity. Using Carr-Purcell-Meiboom-Gill relaxation dispersion experiments, we have measured millisecond dynamics to identify such states. We have also performed concentration dependent relaxation experiments and analytical ultracentrifugation experiments that report on the effective protein size to look for possible differences in self-association for active and autoinhibited forms of the EphB2 kinase domain. We show that the active but not autoinhibited forms exchange between a ground state and an excited state at a rate of 1900 s-1. Similar results were found for the S677/680A mutant of the protein. The nature and importance of the excited state is still unknown. Our most important finding is that active forms of the kinase domain self-associate in a concentration dependent manner and form tetramers and possibly larger oligomers. Multimerization of the kinase domain may enable the assembly of complexes of downstream proteins and could be important for Eph signaling.

Keywords
Kinase activation | Eph receptors | chemical exchange | nmr spectroscopy | protein dynamics | self-association
National Category
Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-117071 (URN)
Available from: 2015-04-15 Created: 2015-04-15 Last updated: 2015-04-15Bibliographically approved
Ahlner, A., Mayzel, M., Lundström, P. & Orekhov, V. Y.Measurement of Protein Backbone 13CO and 15N Relaxation Dispersion at High Resolution.
Open this publication in new window or tab >>Measurement of Protein Backbone 13CO and 15N Relaxation Dispersion at High Resolution
(English)Manuscript (preprint) (Other academic)
Abstract [en]

Three-dimensional pulse sequences for the measurement of Carr-Purcell-Meiboom-Gill relaxation dispersions and new methods for co-processing non-uniformly sampled data are presented. The new methodology was validated for the disordered protein IgA and for an SH3 domain from Abp1p in exchange between its free form and bound to a peptide from the protein Ark1p. We show that the results are similar to ones obtained using traditional experiments and that accurate excited state chemical shifts can be determined. Furthermore, we show that jackknife analysis of down sampled spectra yields robust estimates of peak intensities errors, eliminating the need for recording duplicate data points. The methodology should be useful for characterization of millisecond dynamics in small to medium-sized proteins with poorly dispersed spectra.

National Category
Chemical Sciences
Identifiers
urn:nbn:se:liu:diva-117070 (URN)
Available from: 2015-04-15 Created: 2015-04-15 Last updated: 2015-04-15Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0001-7004-8251

Search in DiVA

Show all publications