Sulphonate buffers affect the recovery of microtubule-associated proteins MAP1 and MAP2: evidence that MAP1A promotes microtubule assembly.

Abstract

The influence of two commonly used sulphonate buffers, PIPES and MES, on the in vitro assembly of bovine brain microtubule protein was examined. Microtubule assembly was monitored by turbimetry and, after centrifugation, the polymerised protein was analysed by SDS-PAGE and western blotting. Assembly in MES when compared with PIPES resulted in a higher recovery of microtubule proteins at both pH 6.4 and pH 6.9 and in an altered protein composition. The buffer pH affected the total amount of protein polymerised but did not significantly affect the protein composition. At both pH conditions the recovery of HMW-MAPs was markedly increased in MES buffer and this increase was mostly due to an increase in the amount of MAP1.