ISSN:
0886-1544
Keywords:
MAP5
;
high-molecular weight MAPs
;
tubulin
;
actin
;
cytoskeleton
;
Life and Medical Sciences
;
Cell & Developmental Biology
Source:
Wiley InterScience Backfile Collection 1832-2000
Topics:
Biology
,
Medicine
Notes:
A simple procedure for the purification of MAP1B from bovine brain is described. The procedure requires two ion-exchange chromatographic steps and results in 〉95% pure MAP1B with a typical recovery of about 25-30 mg/kg of brain tissue. SDS-PAGE analysis of the purified protein shows that it is composed of a high molecular mass (330kDa) heavy chain and two low molecular mass (32kDa and 18kDa) associated light chains. The estimated stoichiometry of heavy chain:light chain is 1:2 and 1:0.2 mole/mole protein for the 32kDa and 18kDa light chains respectively. Western blotting, using monospecific monoclonal antibodies, shows that only the heavy chain is recognised by the anti-MAP1B antibody and is not immunostained by either the MAP1A or MAP2 monoclonal antibodies. Purified MAP1B binds efficiently to both unpolymerised tubulin and polymerised tubulin and co-sediments with taxol-stabilised microtubules. Co-incubation experiments show that MAP2 can compete with MAP1B binding to microtubules, indicating common or overlapping sites. However, MAP1B binds to neither G-actin nor F-actin nor co-sediments with F-actin, suggesting that it is not an actin-binding protein.
Additional Material:
7 Ill.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1002/cm.970300407
