Influence of elastin-like peptide fusions on the quantity and quality of a tobacco-derived human immunodeficiency virus-neutralizing antibody.
The use of vaginal microbicides containing human immunodeficiency virus (HIV)-neutralizing antibodies (nAbs) is a promising strategy to prevent HIV-1 infection. Although antibodies are predominantly manufactured using mammalian cells, elastin-like peptide (ELP) fusion technology improves the stability of recombinant, plant-produced proteins and facilitates their purification, making plants an alternative platform for antibody production. We generated transgenic tobacco plants accumulating four different formats of the anti-HIV-1 antibody 2G12 in the endoplasmic reticulum (ER), i.e. with ELP on either the light or heavy chain, on both, or on neither. Detailed analysis of affinity-purified antibodies by surface plasmon resonance spectroscopy showed that the kinetic binding parameters of all formats were identical to 2G12 lacking ELP produced in Chinese hamster ovary (CHO) cells. Importantly, protein purification from seeds by inverse transition cycling (ITC) did not affect the binding kinetics. Analysis of heavy chain N-glycans from leaf-derived antibodies showed that retrieval to the ER was efficient for all formats. In seeds, however, N-glycans on the naked antibody were extensively trimmed compared with those on the ELP fusion formats, and were localized to a different subcellular compartment. The in vitro HIV-neutralization properties of the tobacco-derived 2G12 were equivalent to or better than those of the CHO counterpart.
Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Gatersleben, Germany.
Doreen M
DM
Floss
Markus
M
Sack
Elsa
E
Arcalis
Johannes
J
Stadlmann
Heribert
H
Quendler
Thomas
T
Rademacher
Eva
E
Stoger
Jürgen
J
Scheller
Rainer
R
Fischer
Udo
U
Conrad
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D057134:Antibodies, Neutralizing / Q000096:biosynthesis* / Q000302:isolation & purification; D000080908:Broadly Neutralizing Antibodies; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004549:Elastin / Q000235:genetics*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D005822:Genetic Vectors; D006031:Glycosylation; D015483:HIV Antibodies / Q000096:biosynthesis* / Q000302:isolation & purification; D009500:Neutralization Tests; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000009:adverse effects* / Q000096:biosynthesis; D012639:Seeds / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism*
C529157:2G12 monoclonal antibody; D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D000080908:Broadly Neutralizing Antibodies; D015483:HIV Antibodies; D011993:Recombinant Fusion Proteins; D004549:Elastin