Protein X-Ray Crystallography

In order to understand how macromolecules, particularly proteins, function — for example how they transport oxygen, detect light, fire an electrical impulse, break down or synthesise other molecules, etc. — we need to know their three-dimensional structures.

Structural information of proteins can also be useful for industrial or medical applications. Structure-based engineering of α-amylases, a well-researched important enzyme of vast industrial value, is still a keen research topic in order to meet the different industrial requirements. Likewise, based on the proteins’ known structures, drugs can efficiently be designed to inhibit proteins which are involved in diseases such as dengue fever and malaria.

X-ray crystallography is the primary means of solving a protein’s structure. The process includes crystallization, crystal optimization, crystal diffraction and structure determination. Since every protein crystallizes under different conditions, over hundreds of conditions have to be screened during crystallization to identify the initial crystallization conditions, which will then need to be optimized. The Protein X-Ray Crystallography unit is fully equipped to carry out structural studies on proteins or protein complexes (with a ligand, DNA, RNA, other protein, etc.).

In order to understand how macromolecules, particularly proteins, function — for example how they transport oxygen, detect light, fire an electrical impulse, break down or synthesise other molecules, etc. — we need to know their three-dimensional structures.

Crystallization Screening
  • Robotic setup in a 96-well plate, with daily inspection for the first week and then weekly for up to one month
  • Available commercial screens including Crystal Screens 1 and 2 (Hampton Research), Structure Screens 1 and 2 (Molecular Dimensions), Wizard I–IV (Emerald BioSystems), Pi-minimal Screen (Jena Bioscience), etc
Crystal Optimization
  • Optimization of the parameters (pH, concentrations, temperature, etc) of promising crystallization conditions
  • Co-crystallization with a ligand if available
Data Collection
  • Diffraction tests for crystal quality checking
  • Determination of cryogenic conditions
  • Collection of a full diffraction data set
Structure Determination
  • Processing of diffraction data
  • Phase determination using molecular replacement or single anomalous diffraction (SAD)
  • Model building and refinement

The Protein X-Ray Crystallography unit houses the following facilities:

  • Rigaku X-ray generator with a copper anode, MicroMax-007 HF, coupled with the VariMax HF optic and R-AXIS IV++ IP area detector. This powerful in-house X-ray diffraction system is ideal for a wide range of protein crystals.
  • Phoenix liquid handling system from Art Robbins. The system can speedily dispense as little as 100 nl of a protein sample into a drop, hence greatly reducing the amount of sample and time needed for setting up crystallization screens.
  • Rigaku UV and visible crystal imaging and protein crystal monitoring system, Minstrel DT UV. It can detect crystals where visible imaging falls short such as in heavily precipitated drops, and can easily distinguish protein crystals from salt crystals.

CCB's Protein X-Ray Crystallography service may vary from weeks to months, according to the differential parameter involved. Thus, please contact us for the discussion of your project. We will ensure a quick service at highly competitive price.

Centre For Chemical Biology (CCB@USM),
1st Floor. Block B, No-10
Persiaran Bukit Jambul,
11900 Bayan Lepas,Penang
Tel: 604-6535502 Fax: 604-6535514
Email: amrinarosyada@usm.my