Identification of Acan125 as a Myosin-I-binding Protein Present with Myosin-I on Cellular Organelles of Acanthamoeba

doi:10.1074/jbc.270.43.25316

Identification of Acan125 as a Myosin-I-binding Protein Present with Myosin-I on Cellular Organelles of Acanthamoeba(*)

From the Department of Physiology, University of Texas Southwestern Medical Center, Dallas, Texas 75235-9040

^§ To whom correspondence should be addressed:
Dept of Physiology, UT Southwestern Medical Center, 5323 Harry Hines Blvd., Dallas, TX 75235-9040.
Tel.: 214-648-7224; Fax: 214-648-8685; zot{at}utsw.swmed.edu.

Abstract

We have discovered the first protein to bind to a nonfilamentous myosin, aside from actin. This protein, Acan125, is a 125-kDa protein from Acanthamoeba that associates with the SH3 domain of Acanthamoeba myosin-IC and not the SH3 domain of human fodrin. Antibodies raised against Acan125 recognize a single protein of 125 kDa from a whole cell lysate of Acanthamoeba; antibodies to myosin-I (M1.7 and M1.8) do not recognize Acan125 on the same blot. Double labeling of Acanthamoeba show Acan125 and myosin-I to be present on the same intracellular organelle, most likely amoebastomes. Immunoprecipitation with either anti-myosin-I or anti-Acan125 antibodies coprecipitates both Acan125 and myosin-I from a lysate of Acanthamoeba, demonstrating that Acan125 interacts with native myosin-I.

Footnotes

↵* This work was supported by Grant MCB-9205344 from the National Science Foundation and Grant 92-1571 from the American Heart Association National Center. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
¹ The abbreviations used are:

SH3

Src homology 3

GST

glutathione S-transferase

TBS

Tris-buffered saline

PAGE

polyacrylamide gel electrophoresis

PVDF

polyvinylidene difluoride.
↵²In a preliminary report (Zot, A. S., and Zot, H. G.(1994) Mol. Biol. Cell5, 162A), Acan125 was incorrectly estimated to be 150 kDa.
↵³S. K. Doberstein, I. C. Baines, E. D. Korn, and T. D. Pollard, personal communication.
- Received June 28, 1995.
- Revision received August 31, 1995.