Protein–Protein Interaction Detection: Methods and Analysis

• Published in: Plant Biotechnology Vol. 1; pp. 391 - 411
• Main Authors: Sharma, Vaishali, Ranjan, Tushar, Kumar, Pankaj, Pal, Awadhesh Kumar, Jha, Vijay Kumar, Sahni, Sangita, Prasad, Bishun Deo
• Format: Book Chapter
• Language: English
• Published: CRC Press 2018
• Edition: 1
• Subjects: Phage Genomic DNA; electrostatic force; High Throughput Screening; Cell Cycle Control; Glutathione Agarose Beads; SPR; Aspartate Transcarbamylase; Fluorescence Resonance Energy Transfer; Protein Fragment Complementation Assay; multisubunit; Filamentous Phage; Stable PPIs; Glycyl tRNA Synthetase; Peptide BDs; leucin zipper; Gene III; GAL4; hydrophobic interaction; Multisubunit Proteins; Y2H; Unknown Proteins; Phage Display Technique; ITC; Fret; Multisubunit Complexes; Phage Display; Core DNA; Protein Interaction Analysis
• ISBN: 9781771885805; 1771885807
• DOI: 10.1201/9781315213743-18

Protein–protein interactions (PPIs) are ubiquitous to cellular process. Proteins are the workhorses that facilitate most biological processes in a cell, including gene expression, DNA replication, transcription, translation, splicing, secretion, cell cycle control, signal transduction, apoptosis, and intermediary metabolism, where PPI plays a crucial role to facilitate their smooth running. Implications about function of any protein can be made via PPI. These implications are based on the premise that the function of unknown proteins may be discovered if captured through their interaction with a protein target of known function. These interactions are critical for the interactomics system of the living cell (or organisms) and abnormal PPIs are the sole reason for several diseases such as Alzheimer’s disease and cancer. Extracellular signal basically regulates the activities of cells. Transduction of signals inside the interior of cells depends critically on PPIs between the various signaling molecules and transmembrane proteins. These signals could be for either normal metabolism or even cell death. Not only this, but also PPIs help in carrying proteins from one place to targets (such as from cytoplasm to nucleus in case of the nuclear importin). In most of the cases, one protein carries another protein for proper functioning of cells. In almost all biosynthetic events in cells, enzymes interact or communicate with each other and generate other macromolecules. Regulation of all metabolic events (e.g., negative feedback) and biology of muscle contraction also involves several interactions. A well-known binding motif that facilitates relatively stable protein–protein interactions (PPIs) is the leucine zipper, comprising a-helices on each protein that binds to each other in a parallel fashion through the hydrophobic interaction. Publication of the human genome and proteomics-based protein profiling studies catalyzed resurgence in protein interaction analysis. Multisubunit proteins are much more efficient in terms of energy utilization relative to a single large protein with multiple sites. For two-hybrid system, the transcription factor is split into two separate domains, known as binding domain and activating domain. Pull-down assays are akin in methodology to coIP because of the use of beaded support to purify interacting proteins. The display beautifully explains a connection between genotype and phenotype. The process of resonance energy transfer can take place when a donor fluorophore in an electronically excited state transfers its excitation energy to a nearby chromophore, the acceptor.