Functional mutations in and characterization of VHH against Helicobacter pylori urease.
Manipulation of clinically significant antibodies can effectively improve the processes of diagnosis and treatment. Affinity maturation process has a significant role in improvement of antibodies efficiency. Error-prone PCR technique is one of the proposed methods for improvement of the affinity of antibodies. In the present research, a method was applied to camel heavy-chain antibody (VHH, nanobody) raised against UreC subunit of urease enzyme from Helicobacter pylori. This VHH was used as a starting molecule to construct a highly diversified phage displayed VHH library. The constructed library of nanobody mutants was subjected to several rounds of panning against UreC antigen. High-affinity mutant was selected. Our VHH (HMR23) showed 1.5-fold higher binding activity than the parental VHH. In addition, the mutant VHH presented a better performance in inhibition of urease activity at low concentrations retaining its specificity and thermal stability.
Department of Biology, Faculty of Basic Science, Shahed University, Opposite Imam Khomeini's Shrine, Tehran-Qom Express way, Tehran, 3319118651, Iran.
Reyhaneh
R
Hoseinpoor
Seyed Latif
SL
Mousavi Gargari
Iraj
I
Rasooli
Masoumeh
M
Rajabibazl
Bahareh
B
Shahi
D000818:Animals; D000915:Antibody Affinity; D000918:Antibody Specificity; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D001483:Base Sequence; D002162:Camelus; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D016480:Helicobacter pylori / Q000737:chemistry* / Q000201:enzymology / Q000276:immunology; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D019151:Peptide Library; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D061905:Single-Domain Antibodies / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D014510:Urease / Q000737:chemistry* / Q000235:genetics / Q000276:immunology
D001426:Bacterial Proteins; D007143:Immunoglobulin Heavy Chains; D019151:Peptide Library; D011994:Recombinant Proteins; D061905:Single-Domain Antibodies; D014510:Urease