Self-catalyzed insertion of proteins into phospholipid membranes

We present evidence that the mitochondrial channel, VDAC, when reconstituted into a phospholipid membrane, can catalyze the insertion of other VDAC channels. This property called "auto-directed insertion" was first proposed by Zizi et al. (1995) to explain observations on asymmetric VDAC channels. We found that 2 urea or guanidinium chloride (GdmCl) caused a burst of insertions of VDAC channels when added to the same side as VDAC addition. More strikingly, when added to the opposite side they caused a 10-60-fold sustained yet reversible increase in insertion rate. Protein stabilization by sarcosine eliminated the effect of urea and GdmCl on VDAC insertion. Control experiments showed that water flow, ionic strength, osmotic force, phospholipid type, and membrane potential were not involved. Therefore, although both urea and GdmCl affect the properties of phospholipid membranes, it is more likely that these agents act either by changing the structure of the pre-inserted channels, allowing them to be more effective catalysts for VDAC insertion, or by flowing through the channels and acting on nearby VDAC channels inducing them to insert. Either way, insertion must be occurring next to pre-inserted channels. Urea and GdmCl may mimic chaperones by partially unfolding VDAC and keeping it in an insertion-competent state. "Auto-directed insertion" may ensure both correct targeting and orientation of nascent proteins in vivo.