Association behaviour of human beta B1-crystallin and its truncated forms

beta B1-crystallin plays an important role in the assembly of betaH-crystal lin yet is known to be subject to N-terminal sequence truncations during hu man lens development and ageing. Here we have over-expressed human beta B1- crystallin, and various truncated forms in Escherichia coli and used mass s pectrometry to monitor the monomer molecular weight. Gel permeation chromat ography and laser light scattering have been used to estimate the assembly size of the various polypeptides as a function of protein concentration. Th e full-length beta BI-crystallin behaves as a dimer, like recombinant human beta B2-crystallin, but undergoes further self-association at high protein concentrations, unlike the beta B2-crystallin, Major truncations from the N-terminat extension lead to anomalous behaviour on gel permeation chromato graphy indicative of altered interactions with the column matrix, whereas l ight scattering indicated dimers at low protein concentration that self-ass ociate as a function of protein concentration. Loss of 41 residues from the N-terminus, equivalent to an in vivo truncation site, resulted in temperat ure-dependent phase separation behaviour of the shortened beta B1-crystalli n. Good crystals have been grown of a truncated version of human beta BI-cr ystallin using an in vitro cleavage protocol. (C) 2001 Academic Press.