A structural insight into the molecular recognition of a (−)-Δ9-tetrahydrocannabinol and the development of a sensitive, one-step, homogeneous immunocomplex-based assay for its detection

Merja H. Niemi, Laura Turunen, Timo Pulli, Tarja Nevanen, Matti Höyhtyä, Hans Söderlund, Juha Rouvinen (Corresponding Author), Kristiina Takkinen

Research output: Contribution to journalArticleScientificpeer-review

5 Citations (Scopus)

Abstract

(−)-Δ9-Tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC–bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 Å and 2.0 Å resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity: the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C10 monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C10 monoterpene moiety, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol and 11-hydroxy-Δ9-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab–Δ9-THC immunocomplex binding antibody from a naïve antibody phage display library, the specificity of the Δ9-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ng/ml from saliva samples.
Original languageEnglish
Pages (from-to)803-814
Number of pages12
JournalJournal of Molecular Biology
Volume400
Issue number4
DOIs
Publication statusPublished - 2010
MoE publication typeA1 Journal article-refereed

Fingerprint

Dronabinol
Monoterpenes
Libraries
Fluorescence Resonance Energy Transfer
Immunoglobulin Fab Fragments
Antibodies
Haptens
Cannabis
Saliva
Immunoassay
Serum Albumin
Bacteriophages
Limit of Detection
Hydrogen
Spleen
Binding Sites

Keywords

  • antibody Fab fragment
  • crystal structure
  • tetrahydrocannabinol
  • hapten binding
  • cannabinoid

Cite this

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title = "A structural insight into the molecular recognition of a (−)-Δ9-tetrahydrocannabinol and the development of a sensitive, one-step, homogeneous immunocomplex-based assay for its detection",
abstract = "(−)-Δ9-Tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC–bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 {\AA} and 2.0 {\AA} resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity: the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C10 monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C10 monoterpene moiety, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol and 11-hydroxy-Δ9-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab–Δ9-THC immunocomplex binding antibody from a na{\"i}ve antibody phage display library, the specificity of the Δ9-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ng/ml from saliva samples.",
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A structural insight into the molecular recognition of a (−)-Δ9-tetrahydrocannabinol and the development of a sensitive, one-step, homogeneous immunocomplex-based assay for its detection. / Niemi, Merja H.; Turunen, Laura; Pulli, Timo; Nevanen, Tarja; Höyhtyä, Matti; Söderlund, Hans; Rouvinen, Juha (Corresponding Author); Takkinen, Kristiina.

In: Journal of Molecular Biology, Vol. 400, No. 4, 2010, p. 803-814.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - A structural insight into the molecular recognition of a (−)-Δ9-tetrahydrocannabinol and the development of a sensitive, one-step, homogeneous immunocomplex-based assay for its detection

AU - Niemi, Merja H.

AU - Turunen, Laura

AU - Pulli, Timo

AU - Nevanen, Tarja

AU - Höyhtyä, Matti

AU - Söderlund, Hans

AU - Rouvinen, Juha

AU - Takkinen, Kristiina

PY - 2010

Y1 - 2010

N2 - (−)-Δ9-Tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC–bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 Å and 2.0 Å resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity: the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C10 monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C10 monoterpene moiety, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol and 11-hydroxy-Δ9-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab–Δ9-THC immunocomplex binding antibody from a naïve antibody phage display library, the specificity of the Δ9-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ng/ml from saliva samples.

AB - (−)-Δ9-Tetrahydrocannabinol (THC) is the main psychoactive compound found in cannabis. In this study, an anti-THC Fab fragment, designed T3, was isolated from a display library cloned from the spleen cells of a mouse immunized with a THC–bovine serum albumin conjugate, and the crystal structures of the T3 Fab in its free form and in complex with THC were determined at 1.9 Å and 2.0 Å resolution, respectively. The THC binding site of the T3 Fab is a narrow cavity: the n-pentyl group of THC protrudes deep into the interface area between the variable domains and the C10 monoterpene moiety of the hapten is partially exposed to solvent. The metabolites of THC, with modifications in the C10 monoterpene moiety, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol and 11-hydroxy-Δ9-tetrahydrocannabinol, are bound by the T3 Fab with a higher affinity than THC. The crystal structures suggest that Ser52H and Arg53H of the T3 Fab are able to make hydrogen bonds with the metabolites, which leads to an increased binding against these metabolites. By developing a T3 Fab–Δ9-THC immunocomplex binding antibody from a naïve antibody phage display library, the specificity of the Δ9-THC binding is highly increased, which allows a one-step, homogeneous, fluorescence resonance energy transfer-based sensitive immunoassay, with a detection limit of 20 ng/ml from saliva samples.

KW - antibody Fab fragment

KW - crystal structure

KW - tetrahydrocannabinol

KW - hapten binding

KW - cannabinoid

U2 - 10.1016/j.jmb.2010.05.048

DO - 10.1016/j.jmb.2010.05.048

M3 - Article

VL - 400

SP - 803

EP - 814

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 4

ER -