Synthetic hypusine peptides

Details Synthetic hypusine peptides Synthetic hypusine peptides

2000-03-07

2002-12-10

Low, Christopher S. F. (Department: 1653)

Chemistry: natural resins or derivatives; peptides or proteins;

Peptides of 3 to 100 amino acid residues

6 to 7 amino acid residues in defined sequence

C530S328000, C530S330000, C514S016700, C514S017400, C562S553000, C562S562000

Reexamination Certificate

active

06492489

ABSTRACT:

BACKGROUND OF THE INVENTION
Hypusine [N
&egr;
-(4-amino-2-hydroxybutyl) lysine], or [2S, 9R)-2, 11-diamino-9-hydroxy-7-azaundecanoic acid], an unusual naturally occurring amino acid, having the structure:
was first isolated from bovine brain extracts by Shiba et al. in 1971
[Biochim. Biophys. Acta.
Vol. 244, pages 523-531 (1971)]. The molecule has two chiral centers at positions 2 and 9, each of which can be classified R or S by the Cahn-Ingold-Prelog method. The (2S, 9R) diastereomer (B), formed as a post-translational modification
of lysine, has been shown to occur on a precursor protein of the eukaryotic initiation factor 5A (formerly called elF-4D) [Cooper et al.
Proc. Natl. Acad. Sci. U.S.A.,
Vol. 80, pages 1854-1857 (1983); and Safer,
Eur. J. Biochem.,
Vol. 186, pages 1-3 (1989)]. This initiation factor 5A is unique in that it is the only known cellular protein that contains the amino acid hypusine (Hpu). In the mid-1970's elF-5A was shown to stimulate ribosomal subunit joining and to enhance 80 S-bound Met-t-RNA
i
reactivity with puromycin [Anderson et al.,
FEBS Lett.,
Vol. 76, pages 1-10 (1977); and Kemper et al.,
J. Biol. Chem.,
Vol. 251, pages 5551-5557 (1976)]. Later, in 1983, Cooper et al., supra, suggested that a hypusine-modified protein serves as an important initiation factor in all growing eukaryotic cells. In 1986, Park et al.,
J. Biol. Chem.,
Vol. 261, pages 14515-14519 (1986)] isolated the elF-5A protein from human red blood cells and elucidated the amino acid sequences surrounding the single hypusine residue, as Thr-Gly-Hpu-His-Gly-His-Ala-Lys (SEQ ID NO: 1). Furthermore, and most interesting because of the potential application to the control of HIV replication [Bevec et al.,
J. Proc. Natl. Acad. Sci. U.S.A.,
Vol. 91, pages 10829-10833 (1994); and Ruhl et al.,
J. Cell Biol.,
Vol. 123, pages 1309-1320 (1994)], the synthesis of elF-5A analogues are of great therapeutic significance.
Since hypusine is specific to elF-5A, antibodies derived from hypusine-containing peptides could be used to quantitate the levels of elF-5A directly and with high specificity. Interest in developing an antibody assay of elF-5A to investigate the physiological role of this important initiation factor prompter total synthesis of hypusine and its (2S, 9R)-diastereomer [Bergeron et al.,
J. Org. Chem.,
Vol. 58, pages 6804-6806 (1993)]. The key step in the synthesis involved the N
&egr;
-alkylation of N
&egr;
-benzyl-N
&agr;
-carbobenzyloxy-(L)-lysine benzyl ester with (R)- or (S)-epichlorohydrin to give the respective (2S, 9R)- and (2S, 9S)-chlorohydrins. Subsequent displacement of the respective chlorides by cyanide ion provided the protected hypusine skeletons. The final step, hydrogenation over PtO
2
in AcOH, followed by neutralization and re-acidification, yielded the respective (2S, 9S)- and (2S, 9R)-hypusine dihydrochlorides. A comparison of the reported hypusine optical rotation with that of the synthetic (2S, 9R)-hypusine B confirmed the stereochemical integrity of both chiral centers throughout the synthesis.
Synthetic methodology for accessing hypusine itself exists and it was desirable to have a selectively-protected hypusine reagent which could be used to incorporate this unusual amino acid into selected peptides. Copending application Ser. No. 08/962,300, filed Oct. 31, 1997 now U.S. Pat. No. 5,973,113 entitled “Hypusine Reagent for Peptide Synthesis,” the entire contents and disclosure of which are incorporated herein by reference, describes a selectively protected hypusine reagent useful for incorporating hypusine into peptides, as well as methods for preparation of the hypusine reagent.
It is an object of the present invention to provide novel hypusine-containing peptides, as well as methods for their synthesis utilizing the above-described hypusine reagent.
SUMMARY OF THE INVENTION
The above and other objects are realized by the present invention, one embodiment of which relates to novel peptides containing the hypusine moiety.
More specifically, the present invention relates to novel hypusine-containing peptides synthesized utilizing the hypusine reagent:
wherein:
Q
1
, Q
2
and Q
3
may be the same or different and are amino protective groups, provided that Q
3
is orthogonal to Q
1
and Q
2
; and
Z is a hydroxy protective group.
A further embodiment of the invention relates to compounds of structure (2) which may be synthesized using hypusine reagent (1), wherein Hpu is the hypusine amino acid residue, S and T are each independently peptide residues from zero to about 12 amino acids in length. Compounds of the invention find utility in the study of biochemical processes involving hypusine.
Another embodiment of the invention concerns improved methods of peptide synthesis wherein the above-described hypusine reagent is employed to prepare novel hypusine-containing peptides.


REFERENCES:
patent: 5344846 (1994-09-01), Jakus et al.
patent: 5538897 (1996-07-01), Yates, III et al.
Bergeron, R.J., et al., “Synthesis of Reagents for the Construction of Hypusine and Deoxyhypusine Peptides and Their Application as Peptidic Antigens,”J. Med. Chem., 41(20):3888-3900 (1998).
Shiba, T., et al., “Hypusine, A New Amino Acid Occurring in Bovine Brain Isolation and Structural Determination,”Biochim. Biophys. Acta, 244(3):523-531 (1971).
Imaoka, N. and Nakajima T., “Hypusine N6—(4-Amino-2-Hydroxybutyl) -2,6-Diaminohexanoic Acid, In Tissue Proteins of Mammals,”Biochem. Biophys. Acta, 320:97-103 (1973).
Park, M. H., et al., “Identification of Hypusine, an Unusual Amino Acid, in a Protein from Human Lymphocytes and of Spermidine as its Biosynthetic Precursor,”Proc. of the Nat. Acad. of Sci., 78(5):2869-2873 (1981).
Sano, A., et al., “Isolation and Identification of &agr;-(&ggr;-Aminobutyryl)—Hypusine,”J. of Neurochemistry, 46(4):1046-1049 (1986).
Park, M. H., et al., “Eukaryotic Initiation Factor 4D Purification From Human Red Blood Cells and the Sequence of Amino Acids Around its Single Hypusine Residue,”J. of Bio. Chem., 261(31):14515-14519 (1986).
Sandholzer, U., et al., “cDNA and Derived Amino Acid Sequence of the Hypusine Containing Protein fromDictyostelium discoideum,”FEBS Letters, 246, Issue 1-2:94-100 (1989).
Beninati, S., et al., “High-Performance Liquid Chromatographic Method for Determination of Hypusine and Deoxyhypusine,”Analytical Biochemistry 184(1):16-20 (1990).
Ueno, S., et al., “Isolation and Identification of &agr;- (&bgr;-alanyl) Hypusine from Bovine Brain,”Biochemica et Biophysica Acta, 1073(1):233-235 (1991).
Bergeron,R.J. et.al., “Total Syntheses of (+)-Hypusine and Its (2S,9S)—Diastereomer,”J. Org. Chem. 58:6804-6806 (1993).
Beyermann, M. et al., “Rapid Continuous Peptide Synthesis via FMOC Amino Acid Chloride Coupling and 4—(Aminomethyl)piperidine Deblocking,”J. Org. Chem., 55(2):721-728 (1990).
Martinez, J. et al., “On the Use of Carboxamidomethyl Esters (CAM Esters) in the Synthesis of Model Peptides. Scope and Limitations,”Tetrahedron, 41(4):739-743 (1985).
Martinez, J. et al., “Carboxamidomethyl Esters (CAM Esters) as Carboxyl Protecting Groups,”Tetrahedron Letter., 24(47):5219-5222 (1983).
Moroder, L. et al., “A New Synthetic Route to Tyrosine-O-Sulfated Peptides: Synthesis of a Pancreaozymin-Caerulein-Hybrid Related Peptide,” inHormone Receptors in Digestion and Nutrition, G. Rosselin, et al. eds., Elsevier/North Holland Biomedical Press, Amsterdam, 129-135 (1979).
Moroder, L. et al., “Synthese von Tyrosin-O-sulfat-haltigen Peptiden,”Z. Physiol. Chem., 360:787-790 (1979).
Shioiri, T. et al., “Diphenylphosphoryl Azide. A New Convenient Reagent for a Modification Curtius Reaction and for the Peptide Synthesis,”J. Am. Chem. Soc., 94(17):6203-6205 (1972).
Wang, S.S. et al. “Cleavage and deprotection of peptides on MBHA-resin with hydrogen bromide,”Int. J. Peptide Protein Res., 40:344-349 (1992).
Yamashiro, D. et al., “The Use of N&agr;, Nim-Bis (tert-butyloxycarbonyl) Histidine and N&agr;-2-(p-Biphenylyl)isopropyloxycarbonyl-Nim-tert-butyloxy-carbony

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