Should I always removeEventListener?

Should I then remove the load event listener from the window after the event is fired?

I've never seen that done, so I don't think there's a real need for it.

It only fires once, but will it continue to listen after that happens?

It is only fired once by the DOM, yes. But it will continue to listen, you could easily trigger a load event manually (see MDN for examples).

But is that overkill or will that actually benefit the performance of my program?

Typically it's overkill, as this doesn't make a huge difference. Of course, it might trigger garbage collection on initialize, which could save a bit of memory (or more, depending on your code structure) and improve performance by making it available to the rest of your app.

window.addEventListener('load', initialize, {once: true});