HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Yamaguchi, T. Articles by Adachi, K. Search for Related Content PubMed PubMed Citation Articles by Yamaguchi, T. Articles by Adachi, K. Social Bookmarking                      What's this?

Volume 271, Number 43, Issue of October 25, 1996 pp. 26677-26683
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Expression of Soluble Human -Globin Chains in Bacteria and Assembly in Vitro with -Globin Chains

(Received for publication, July 12, 1996, and in revised form, August 9, 1996)

Takamasa Yamaguchi , Jian Pang , Konda S. Reddy ^§ , H. Ewa Witkowska ^¶ , Saul Surrey and Kazuhiko Adachi

From  The Children's Hospital of Philadelphia, Division of Hematology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104, the ^§ Department of Biophysics, University of Pennsylvania, Philadelphia, Pennsylvania 19104, and the ^¶ Children's Hospital Oakland Research Institute, Oakland, California 94609

Authentic soluble human -globin chains were produced in Escherichia coli using an expression plasmid (pHE2) containing full-length cDNAs coding for human -globin chain and methionine aminopeptidase. Spectral properties of the purified -globin were identical to those of authentic -globin. Soluble -globin showed low (16 kDa) and high molecular mass (32 kDa) forms that could be separated by gel filtration chromatography. SDS-polyacrylamide gel electrophoresis and electrospray mass spectrometry revealed the 32-kDa species was dimeric -globin formed by an intermolecular disulfide bond, while the 16-kDa species was authentic monomeric -globin. Monomeric forms of -globin, like authentic native -globin, formed tetrameric hemoglobin (Hb) A (₂₂) in vitro upon incubation with -globin, while dimeric forms did not. When -globin dimers, however, were converted to monomers by incubation with dithiothreitol, the -globin chain monomers assembled with -globin and formed hemoglobin tetramers. -Globin was more thermally unstable than -globin, while assembled tetramers promoted higher stability. Disulfide-bonded -globin dimers showed a slight increase in thermal stability compared with -globin; however, dimers were still more unstable than tetrameric Hb A. These results indicate that presence of  chains favors assembly with -globin, - dimers cannot bind  chains, and that Hb A tetramer formation results in the most thermally stable species.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				T. Yamaguchi, J. Pang, K. S. Reddy, S. Surrey, and K. Adachi Role of beta 112 Cys (G14) in Homo- (beta 4) and Hetero- (alpha 2beta 2) Tetramer Hemoglobin Formation J. Biol. Chem., June 5, 1998; 273(23): 14179 - 14185. [Abstract] [Full Text] [PDF]

				K. Adachi, T. Yamaguchi, J. Pang, and S. Surrey Effects of Increased Anionic Charge in the beta -Globin Chain on Assembly of Hemoglobin In Vitro Blood, February 15, 1998; 91(4): 1438 - 1445. [Abstract] [Full Text] [PDF]

				K. Adachi, Y. Zhao, T. Yamaguchi, and S. Surrey Assembly of gamma - with alpha -Globin Chains to Form Human Fetal Hemoglobin in Vitro and in Vivo J. Biol. Chem., April 21, 2000; 275(17): 12424 - 12429. [Abstract] [Full Text] [PDF]