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

J. Biol. Chem., Vol. 279, Issue 26, 27518-27524, June 25, 2004

This Article Full Text Full Text (PDF) All Versions of this Article: 279/26/27518    most recent M402578200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted 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 Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Levasseur, D. N. Articles by Townes, T. M. Search for Related Content PubMed PubMed Citation Articles by Levasseur, D. N. Articles by Townes, T. M. Social Bookmarking                      What's this?

A Recombinant Human Hemoglobin with Anti-sickling Properties Greater than Fetal Hemoglobin^*

Dana N. Levasseur¶, Thomas M. Ryan, Michael P. Reilly||**, Steven L. McCune, Toshio Asakura||, and Tim M. Townes

From the Department of Biochemistry and Molecular Genetics, Schools of Medicine and Dentistry, University of Alabama at Birmingham, Birmingham, Alabama 35294 and the ^||Department of Pediatrics and Department of Biochemistry and Biophysics, The Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania 19104

A new recombinant, human anti-sickling -globin polypeptide designated ^AS3 (Gly¹⁶ Asp/Glu²² Ala/Thr⁸⁷ Gln) was designed to increase affinity for -globin. The amino acid substitutions at 22 and 87 are located at axial and lateral contacts of the sickle hemoglobin (HbS) polymers and strongly inhibit deoxy-HbS polymerization. The 16 substitution confers the recombinant -globin subunit (^AS3) with a competitive advantage over ^S for interaction with the -globin polypeptide. Transgenic mouse lines that synthesize high levels of HbAS3 (₂^AS3₂) were established, and recombinant HbAS3 was purified from hemolysates and then characterized. HbAS3 binds oxygen cooperatively and has an oxygen affinity that is comparable with fetal hemoglobin. Delay time experiments demonstrate that HbAS3 is a potent inhibitor of HbS polymerization. Subunit competition studies confirm that ^AS3 has a distinct advantage over ^S for dimerization with -globin. When equal amounts of ^S- and ^AS3-globin monomers compete for limiting -globin chains up to 82% of the tetramers formed is HbAS3. Knock-out transgenic mice that express exclusively human HbAS3 were produced. When these mice were bred with knock-out transgenic sickle mice the ^AS3 polypeptides corrected all hematological parameters and organ pathology associated with the disease. Expression of ^AS3-globin should effectively lower the concentration of HbS in erythrocytes of patients with sickle cell disease, especially in the 30% percent of these individuals who coinherit -thalassemia. Therefore, constructs expressing the ^AS3-globin gene may be suitable for future clinical trials for sickle cell disease.

Received for publication, March 8, 2004 , and in revised form, April 8, 2004.

^* This work was supported in part by National Institutes of Health Grants HL 57619, HL 43508, HL 35559, and HL 38632 and by a grant from the Mizuno Fund. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

These authors contributed equally to this work.

^¶ Supported by National Institutes of Health Predoctoral Fellowship Training Grant 5 T32 CA09467.

^** Current address: Thomas Jefferson University, Philadelphia, PA 19104.

Current address: Northwest Georgia Oncology Centers, P.C., 55 Whitcher St., Ste. 300, Marietta, GA 30060.

To whom correspondence should be addressed: University of Alabama at Birmingham, Dept. of Biochemistry and Molecular Genetics, 502 Kaul Genetics Bldg., 720 20th St. South, Birmingham, AL 35294. Tel.: 205-934-5294; Fax: 205-934-2889; E-mail: ttownes{at}uab.edu.

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

This article has been cited by other articles:

				M. N. Kierlin-Duncan and B. A. Sullenger Using 5'-PTMs to repair mutant beta-globin transcripts RNA, August 1, 2007; 13(8): 1317 - 1327. [Abstract] [Full Text] [PDF]

				J. C. Cheng, K. M. Sakamoto, E. M. Horwitz, S. L. Karsten, L. Shoemaker, H. I. Kornblumc, and P. Malik Report on the Workshop "New Technologies in Stem Cell Research," Society for Pediatric Research, San Francisco, California, April 29, 2006 Stem Cells, April 1, 2007; 25(4): 1070 - 1088. [Abstract] [Full Text] [PDF]