In both my graduate work and during my postodoctoral work I’ve taken advantage of the strong interaction between streptavidin and biotin. Recall the last post we talked about how molecules can fit together into different positions to help stabilize each other. We often think of these in terms of how fast they switch positions from a together state “bound” to an “unbound” state. I’d like to expand upon more complicated systems in later posts but for now we’ll focus on this type of “two state” system. Biotin can position itself inside a small pocket in streptavidin and it loves to do so. Its ability to fit in or bind is known as the binding affinity and is often represented as its dissociation ability or dissociation constant (Kd). The Kd for biotin is about 1*10^-14. That means that for roughly every 100,000,000,000,000 biotin that get into the streptavidin binding pocket only 1 stays out. This makes it almost as strong as a chemical bond! One can do lots of amazing things with bonds these strong but they can also be very annoying.
On the positive side, the bond can be used for biochemical assays. In talking to me you may know that I developed an assay based on streptavidin and biotin binding (or in that case, biotin on a large dextran molecule). The streptavidin-biotin bond is strong enough that even small amounts of the molecules are able to find each other and bind together. This allows us to determine their ability to pass through various barriers like, for instance, a lipid membrane. The binds are tight and will hold together, which gives us a very reliable signal even in different conditions such as very acidic or high salt conditions. Though streptavidin and biotin can be used to work together for some assays, I’m not always happy when they stick together too much.
On the negative side, the binding between Streptavidin and Biotin can pose problems for other scientists. Currently I’m developing an assay to detect target molecules labeled with biotin. The idea is to use biotin to attach the molecule to a streptavidin surface because, again, it is a strong bond. The problem is that my samples might not be pure target-biotin and are contaminated with biotin. Because Streptavidin-biotin has a lower Kd than Streptavidin-target-biotin, the biotin fills the streptavidin pockets before the target-biotin. Also, if I use too little, my molecules no longer bind to the target, they bind to the streptavidin themselves! In this case it’s a bit of a vicious cycle of molecules not fitting into the correct spots! This is why I say that for some applications streptavidin-biotin are great but for others they make a bit of a sticky mess! As always, thanks for reading and come back to enjoy future posts talking about heat and kD and Molecular Yoga!