Noncationic peptides obtained from azurin preferentially enter cancer cells.
Azurin, a member of the cupredoxin family of copper containing redox proteins, preferentially penetrates human cancer cells and exerts cytostatic and cytotoxic (apoptotic) effects with no apparent activity on normal cells. Amino acids 50 to 77 (p28) of azurin seem responsible for cellular penetration and at least part of the antiproliferative, proapoptotic activity of azurin against a number of solid tumor cell lines. We show by confocal microscopy and fluorescence-activated cell sorting that amino acids 50 to 67 (p18) are a minimal motif (protein transduction domain) responsible for the preferential entry of azurin into human cancer cells. A combination of inhibitors that interfere with discrete steps of the endocytotic process and antibodies for caveolae and Golgi-mediated transport revealed that these amphipathic, alpha-helical peptides are unique. Unlike the cationic cell-penetrating peptides, alpha-helical antennapedia-like, or VP22 type peptides, p18 and p28 are not bound by cell membrane glycosaminoglycans and preferentially penetrate cancer cells via endocytotic, caveosome-directed, and caveosome-independent pathways. Once internalized, p28, but not p18, inhibits cancer cell proliferation initially through a cytostatic mechanism. These observations suggest the azurin fragments, p18 and p28, account for the preferential entry of azurin into human cancer cells and a significant amount of the antiproliferative activity of azurin on human cancer cells, respectively.
Taylor
Brad N
BN
Department of Surgical Oncology, University of Illinois College of Medicine, Chicago, Illinois, USA.
Mehta
Rajeshwari R
RR
Yamada
Tohru
T
Lekmine
Fatima
F
Christov
Konstantin
K
Chakrabarty
Ananda M
AM
Green
Albert
A
Bratescu
Laura
L
Shilkaitis
Anne
A
Beattie
Craig W
CW
Das Gupta
Tapas K
TK
D000595:Amino Acid Sequence; D001400:Azurin / Q000493:pharmacokinetics* / Q000494:pharmacology; D048708:Cell Growth Processes / Q000187:drug effects; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D045325:HCT116 Cells; D006801:Humans; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D010446:Peptide Fragments / Q000493:pharmacokinetics* / Q000494:pharmacology; D017434:Protein Structure, Tertiary
D010446:Peptide Fragments; D001400:Azurin