Question

I am very new to protein crystallography and visualization. I have been asked to take photos through an optical microscope of the lysozyme powder directly as-is from the supplier. I suspended some of the powder in both paraffin and glycerol (10 mg/mL & 20 mg/mL for each) to observe the crystal form without solubilizing it.

I'm not sure what it is I'm looking at, however, in all of the suspensions I prepared, the particles look very different from literature images of both lysozyme and salt crystals. The image I included is a 10 mg/mL suspension in paraffin but is essentially what I'm seeing in all cases. A mixture of large chunks with some smaller flakes.

I am wondering if anyone with protein crystal experience can tell me what this form I'm seeing is? And perhaps any further suggestions on how to visualize the sample without any alteration

Answer 1

Lysozyme powder observed under an optical microscope may not show well-defined crystals as it might be partly amorphous. For proper visualization of the protein's structure, X-ray crystallography, not optical microscopy, is typically used to obtain detailed interference patterns indicative of the atomic structure.

Step-by-step explanation:

You are observing the lysozyme powder using an optical microscope to understand the crystal form without solubilizing it. What you are encountering—a mixture of large chunks and smaller flakes—may not resemble typical lysozyme crystals or salt crystals because the powder may not have formed into well-defined crystals, or it could be in a partly amorphous state. High-quality images of protein structures, including enzymes and other challenging systems like membrane-bound receptors, are often obtained through X-ray crystallography, which differs from optical microscopy. X-ray crystallography provides interference patterns that can reveal intricate structural details at the atomic level, which optical microscopy cannot.

It's important to know that lysozyme, as received from suppliers, may not always be in crystalline form suitable for direct microscopic visualization. Suspensions in paraffin or glycerol may also influence the appearance of the particles, as these mediums can affect their dispersal and how they settle on the microscope slide. You may consider ensuring that the sample is properly dispersed and free from aggregates before imaging. If the goal is to visualize the structure of lysozyme, it is recommended to grow dedicated lysozyme crystals under controlled conditions that promote crystal formation, then image them using X-ray crystallography.