2D infrared (IR) spectroscopy is a cutting-edge technique, with applications in subjects as diverse as the energy sciences, biophysics and physical chemistry. This book introduces the essential concepts of 2D IR spectroscopy step-by-step to build an intuitive and in-depth understanding of the method. This unique book introduces the mathematical formalism in a simple manner, examines the design considerations for implementing the methods in the laboratory, and contains working computer code to simulate 2D IR spectra and exercises to illustrate involved concepts. Readers will learn how to accurately interpret 2D IR spectra, design their own spectrometer and invent their own pulse sequences. It is an excellent starting point for graduate students and researchers new to this exciting field. Computer codes and answers to the exercises can be downloaded from the authors' website, available at www.cambridge.org/9781107000056.

Temperature-jump (T-jump) two-dimensional infrared spectroscopy (2D IR) is developed, characterized, and applied to the study of protein folding and association. In solution, protein conformational changes span a wide range of timescale from nanoseconds to minutes. Ultrafast 2D IR spectroscopy measures time-dependent structural changes within the protein ensemble by probing the frequency changes associated with amide I backbone vibrations. Combining 2D IR with a perturbing laser-induced T-jump enables the study of conformational dynamics from 5 ns to 50 ms. To access a finer time-sampling of the conformational evolution, a one-dimensional variant of 2D IR, heterodyne-detected dispersed vibrational echo spectroscopy (HDVE), is implemented. The framework for interpreting transient HDVE and 2D IR spectra is developed, and we propose a method to remove the linear absorption distortions along both frequency axes. We first present the T-jump 2D IR spectra of a dipeptide to reveal the general amide I baseline response expected in the absence of conformational change. To facilitate the analysis of T-jump data, singular value decomposition (SVD) is employed for reducing noise, identifying the number of distinguishable states, and separating spectral changes based on shared timescales. Finally, T-jump 2D IR spectroscopy is applied to study the unfolding of ubiquitin, disordering of the 12-residue p-hairpin peptide trpzip2 (TZ2), and the dissociation of insulin dimers to monomers. Experimental results for ubiquitin highlight the importance of linear absorption corrections for interpretation of the data. In response to the T-jump, 2D IR results indicate p-sheet structure melts in ubiquitin with a small amplitude (~10 gs) and large amplitude (17 ms) response. Isotope-labeling T-jump experiments on TZ2 allow for the proposal of a free energy surface in which transitions from a native and misfolded state proceed through a disordered hub-like state with a 1-2 gs timescale. Multiple timescales are observed in the T-jump induced dissociation of insulin. Based on their spectral features and concentration dependence, the insulin timescales can be assigned to dissociation, disordering, and oligomerization processes. With these applications, we demonstrate the capability of T-jump 2D IR spectroscopy to reveal detailed molecular dynamics.

The series Topics in Current Chemistry Collections presents critical reviews from the journal Topics in Current Chemistry organized in topical volumes. The scope of coverage is all areas of chemical science including the interfaces with related disciplines such as biology, medicine and materials science. The goal of each thematic volume is to give the non-specialist reader, whether in academia or industry, a comprehensive insight into an area where new research is emerging which is of interest to a larger scientific audience. Each review within the volume critically surveys one aspect of that topic and places it within the context of the volume as a whole. The most significant developments of the last 5 to 10 years are presented using selected examples to illustrate the principles discussed. The coverage is not intended to be an exhaustive summary of the field or include large quantities of data, but should rather be conceptual, concentrating on the methodological thinking that will allow the non-specialist reader to understand the information presented. Contributions also offer an outlook on potential future developments in the field.

In this thesis, dynamics experiments are developed that can be used to study protein conformational changes such as folding and binding. Every functional motion of a protein is inextricably linked to conformational dynamics. However, most of our insight into protein folding and binding is indirectly obtained through kinetics experiments that measure reaction rates and reveal how fast populations of stable states interconvert. Two-dimensional infrared spectroscopy (2D IR) is the central tool developed in this thesis for protein dynamics experiments due to its combination of time and structural resolution. As a vibrational spectroscopy, 2D IR potentially offers femtosecond time resolution. Its advantages over linear, absorption spectroscopy come through correlating excitation and emission frequencies to allow for a separation of homogenous and inhomogeneous line shape components, and to give rise to structurally sensitive cross-peaks. One general problem was repeatedly addressed in this thesis: how can 2D IR spectra best be modeled to reveal atomistic structural information? The key feature that now sets 2D IR apart from other fast protein probes is that the data can readily be calculated from an atomistic structure or molecular dynamics simulation using the methods developed in this thesis work. Demonstrative applications are presented for the amide 1-11 spectroscopy of NMA, the amide 1'-II' spectroscopy of poly-L-lysine, isotope-edited 2D IR spectroscopy of trpzip2, and transient 2D JR spectroscopy of ubiquitin unfolding after a temperature jump. The emerging paradigm is to interpret 2D IR spectra with the aid of an atomistic, molecular dynamics simulation. The applications to protein binding use the monomer-dimer transition of insulin as a model system. Using a combination of experiments and simulations, this equilibrium was characterized as a function of protein concentration, temperature, and solvent. Finally, as a complement to the structural information provided by 2D IR, dye-labeling and intrinsic tyrosine fluorescence experiments on insulin are described.

Two-Dimensional Optical Spectroscopy discusses the principles and applications of newly emerging two-dimensional vibrational and optical spectroscopy techniques. It provides a detailed account of basic theory required for an understanding of two-dimensional vibrational and electronic spectroscopy. It also bridges the gap between the formal developm