AlphaFold for experimentalists: a field guide
Structure prediction can now answer questions that used to take a crystallographer years. It can also hand you a beautiful, confident, wrong picture. This guide is about telling the difference.
Being finalized The chapters are written and are being checked before publication. Every check in the guide comes with the program that runs it, the command to type, and real output from an example you can download and reproduce line for line. If you would like to hear when the guide is out, send me a one-line email and I will let you know. No mailing list, no spam.
Who it is for
You run experiments. You have a protein, or two proteins you suspect of binding each other, and you want to know what a predicted structure can honestly tell you before you spend six months at the bench. You do not need to program, and you do not need a background in structural biology.
What is in it
- What a prediction is, and what it is not. The confidence measures, pLDDT and PAE, explained in plain terms, and what each one is actually confident about.
- Asking a good question. Choosing constructs, the price you pay for truncating a protein, and how many copies of each chain to include.
- Reading a predicted complex. Interface confidence, the error between chains, and why a real interface looks the same across all five models.
- The traps. Spurious interfaces and cavities at the cut ends of truncated proteins. Low-confidence regions that only look like structure. Chains that touch without being confidently placed. A distant resemblance in shape mistaken for a shared function.
- Controls. How to build a small panel of known binders and known non-binders and run it through exactly the same settings as your question, so that a negative result means something.
- From prediction to experiment. Which mutants to make, and what to test first.
- Checklists and worked examples from publicly available data.
Why I wrote it
My lab studies how parasite proteins take over host cells, which means we spend a great deal of time asking whether protein A binds protein B. Over the past few years I have watched structure prediction change how we work, and I have also watched it mislead us, more than once, in ways that were obvious only in hindsight. Every trap in this guide is one that caught somebody I know.