Cloning, expression, and purification of HIV-2 gp125: A target for HIV vaccination.
The envelope of the human immunodeficiency virus (HIV) is the main target for neutralizing antibodies. We report the cloning, purification, and characterization of two recombinant forms of the envelope glycoprotein gp125 from a primary HIV-2SBL-6669 isolate. Both constructs were truncated at the N- and C-termini, and in the gp125deltav1v2 construct the variable V1 and V2 loops were deleted. The recombinant glycoproteins were stably expressed in Chinese hamster ovarian cells, producing soluble gp125 and gp125deltav1v2 at molecular weights of 74.2 and 56.9 kDa, respectively, and were purified from cell culture supernatants in a single step using Galanthus nivalis lectin chromatography. Circular dichroism analysis indicated a similar secondary structure for gp125 and gp125deltav1v2, and both proteins were recognized by HIV-2 serum antibodies in surface plasmon resonance assays. The high yield and purity of these constructs makes them suitable for structural and functional analyses, as well as vaccine studies.
Sourial
Samer
S
Microbiology and Tumor Biology Center, Karolinska Institutet, S-171 77, Stockholm, Sweden. samer.sourial@mtc.ki.se
Wärnmark
Anette
A
Nilsson
Charlotta
C
Björling
Ewa
E
Achour
Adnane
A
Harris
Robert A
RA
D016915:AIDS Vaccines; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D006224:Cricetinae; D003412:Cricetulus; D015686:Gene Products, env / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology*; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D017384:Sequence Deletion; D054299:env Gene Products, Human Immunodeficiency Virus
D016915:AIDS Vaccines; D015686:Gene Products, env; D011498:Protein Precursors; D011994:Recombinant Proteins; D054299:env Gene Products, Human Immunodeficiency Virus; C066316:gp125 envelope protein, Human immunodeficiency virus 2