Purification and characterization of a mammalian myosin I

Barbara Barylko, Mark C. Wagner, Ofer Reizes, Joseph P. Albanesi

Research output: Contribution to journalArticle

75 Citations (Scopus)

Abstract

Myosin I, an actin-dependent force-generating enzyme, has been purified from three mammalian sources: bovine adrenal medulla, adrenal cortex, and brain. The purification procedure includes extraction of tissue with ATP at low ionic strength and coprecipitation with actin, followed by gel filtration on Sepharose 4B, anion-exchange chromatography on Q Sepharose, and affinity chromatography on ATP-agarose. Mammalian myosin I molecules are composed of a heavy chain of 116 kDa and multiple low molecular weight polypeptides identified as calmodulin. The structural and enzymatic properties of adrenal medulla myosin I were further characterized. This enzyme exhibits high K+,EDTA- and Ca2+-ATPase specific activities (about 0.2 μmol·min-1 per mg of protein), whereas the Mg2+-ATPase activity is very low (1-3 nmol·min-1·mg-1). The Mg2+-ATPase of medulla myosin I is activated by F-actin in a Ca2+-dependent manner: activity is stimulated 40-fold in the presence of EGTA and 90-fold in the presence of 10 μM Ca2+. Two structural domains of the myosin I heavy chain were identified. A 74-kDa chymotryptic fragment contains the catalytic site, while a 36-kDa polypeptide contains the calmodulin-binding sites. These results indicate that mammalian myosin I is more closely related to myosin I from the avian intestinal brush border than to the enzymes isolated from the protozoans Acanthamoeba and Dictyostelium.

Original languageEnglish (US)
Pages (from-to)490-494
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume89
Issue number2
StatePublished - 1992

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Myosin Type I
Actins
Ca(2+) Mg(2+)-ATPase
Adrenal Medulla
Calmodulin
Enzymes
Acanthamoeba
Agarose Chromatography
Peptides
Dictyostelium
Calcium-Transporting ATPases
Myosin Heavy Chains
Egtazic Acid
Adrenal Cortex
Microvilli
Affinity Chromatography
Sepharose
Osmolar Concentration
Gel Chromatography
Anions

ASJC Scopus subject areas

  • Genetics
  • General

Cite this

Purification and characterization of a mammalian myosin I. / Barylko, Barbara; Wagner, Mark C.; Reizes, Ofer; Albanesi, Joseph P.

In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 89, No. 2, 1992, p. 490-494.

Research output: Contribution to journalArticle

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