Structure
Structural biology is an integral staple of our ecosystem. We maintain a carefully curated set of analytical tools to provide a range of services in crystallography, cryogenic electron microscopy, and mass spectrometry. Our approach to solving structural problems begins with a strong foundation in understanding the molecular mechanisms affecting structure interaction – a powerful asset in deducing problems in protein design and drug discovery.
X-ray Crystallography
Our lab features a Formulatrix Rock 1000 protein crystallization imager and Formulatrix NT8 crystallization automation robot. 1500 LCP plate capacity and an array of services for structure determination, read more below.
CryoTEM
Our Titan and Glacios TEM are fundamental tools in our ability to execute structure determination of proteins and other molecules. Our services in cryo include grid optimization, data acquisition, and model construction. Read more in the CryoTEM section below.
Mass Spectrometry
Ichor has access to an extensive suite of high resolution and sensitive spectrometry tools for precise structure elucidation and sample analysis. Read more about what we offer below.
Capabilities
X-ray Crystallography
(Can also be referred to as Primary Crystal Screening): Preliminary conditions for crystallization of a protein are identified using sparse-matrix screens. These screens vary physicochemical parameters, like concentrations of salt, concentrations of precipitant and pH values, and have been designed using knowledge of conditions that have been successful in producing crystals of protein in the literature.
Screens: The physicochemical parameters for crystallization identified from primary crystal screening are varied over a narrow range. This is used to optimize conditions to obtain crystals that can be used for x-ray diffraction experiments.
Co-crystallization involves crystallizing a protein in the presence of a small molecule that binds to the protein. This small molecule could be a substrate, a ligand or a drug that binds to the protein of interest.
Co-crystallization involves crystallizing a protein in the presence of a small molecule that binds to the protein. This small molecule could be a substrate, a ligand or a drug that binds to the protein of interest.
Protein crystals obtained from screening experiments are frozen in liquid nitrogen and exposed to x-rays at synchrotron sources. The data obtained by diffraction of x-rays by the protein crystals are collected.
(structure solution): X-ray diffraction data collected at synchrotron sources are processed to obtain an electron density map. The electron density map in conjunction with the known protein sequence is used to obtain a three-dimensional structure of the protein.
The three-dimensional structures determined by x-ray diffraction are analyzed to obtain information about inter-residue contacts. The structures can also be analyzed to identify conformations changes that might occur upon binding to another protein or a ligand.
Fragment-based screening involves analyzing the binding of fragments of small molecule (110-250 Da) to proteins of interest, either by ligand soaking or co-crystallization, using x-ray diffraction. The binding site of the fragment to the protein can be identified from the resulting electron density maps.
CryoTEM
Proteins loaded onto Cryo-EM grids will be analyzed to determine properties including ice thickness, orientational freedom, and particle size, density, distribution and shape. Process will be repeated and refined to a point of suitable automatic data collection by varying run parameters.
High resolution data will be collected at high speeds of up to 240 movies per hour, including grid insertion, imaging condition setup, suitable data area selection, and active quality control.
Data collected will be subject to analysis and subsequent structure determination as outlined by Ichor. Image processing for motion correction, estimating CTF, particle selection, and 2D classification. 3D structure determination and modeling using reference map, 3D classification, data interpretation and validation, followed by model genesis.
Mass Spectrometry
Hydrogen/deuterium exchange mass spectrometry is an approach to mass spectrometry involving the rapid conversion of amide hydrogens to deuterium isotope when exposed to heavy water (D2O). This method can be used to observe conformational changes and monitor protein dynamics to map ordered and disordered peptide regions.
- Tandem Mass Tag Labeling
- Targeted Proteomics by Parallel-Reaction Monitoring (PRM)
- Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)
- Label-Free Quantitation
- Protein Profiling
- Peptide Mapping
- Protein Target Identification
- Intact Mass Measurement
- Protein sequencing
- Analysis of Glycopeptides and Determination of the Site Glycosylation
- Acetylation, Methylation, Ubiquitination of Nuclear Proteins
- Glycosylation Profiling
- Phosphorylation Site Identification
- ‘Omics Screening Services
- Metabolite Ratio Determination
- Lipid Profiling
- Biomarker Discovery and Validation
