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Identification of the minimal binding region of a Plasmodium falciparum IgM binding PfEMP1 domain.

Semblat JP, Ghumra A, Czajkowsky DM, Wallis R, Mitchell DA, Raza A, Rowe JA - Mol. Biochem. Parasitol. (2015)

Bottom Line: Here we studied an IgM binding domain from a rosette-mediating PfEMP1 variant, DBL4ζ of TM284var1, and found that the minimal IgM binding region mapped to the central region of the DBL domain, comprising all of subdomain 2 and adjoining parts of subdomains 1 and 3.Site-directed mutagenesis of charged amino acids within subdomain 2, predicted by molecular modelling to form the IgM binding site, showed no marked effect on IgM binding properties.Further work is needed to identify the specific interaction site for IgM within the minimal binding region of PfEMP1.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology and Infection Research, Centre for Immunity, Infection and Evolution, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3FL, United Kingdom.

No MeSH data available.


Related in: MedlinePlus

Identification of the minimal IgM binding region of the TM284var1 DBL4ζ domain.(A) Diagram showing the domain composition of the TM284var1 PfEMP1 variant, with the IgM binding DBL4ζ domain underlined in red. (B) Diagram showing the amino acid domain boundaries and IgM binding properties of each DBL4ζ deletion construct. The full length DBL4ζ domain (top bar) contains 16 cysteines as visualized by the dashed lines (construct used in previous work [2]). Seven deletion constructs were made spanning various regions of DBL4ζ. Proteins that bind human IgM are shown as black bars, and non-binding proteins are shown as white bars. (C) The minimal binding region of DBL4ζ (red). Subdomain (SD) 1 (yellow), 2 (blue) and 3 (grey) are shown, and cysteine residues are highlighted by arrowheads. The five charged residues within subdomain 2 predicted to be involved in IgM binding are shown in bold.
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fig0005: Identification of the minimal IgM binding region of the TM284var1 DBL4ζ domain.(A) Diagram showing the domain composition of the TM284var1 PfEMP1 variant, with the IgM binding DBL4ζ domain underlined in red. (B) Diagram showing the amino acid domain boundaries and IgM binding properties of each DBL4ζ deletion construct. The full length DBL4ζ domain (top bar) contains 16 cysteines as visualized by the dashed lines (construct used in previous work [2]). Seven deletion constructs were made spanning various regions of DBL4ζ. Proteins that bind human IgM are shown as black bars, and non-binding proteins are shown as white bars. (C) The minimal binding region of DBL4ζ (red). Subdomain (SD) 1 (yellow), 2 (blue) and 3 (grey) are shown, and cysteine residues are highlighted by arrowheads. The five charged residues within subdomain 2 predicted to be involved in IgM binding are shown in bold.

Mentions: Previously we studied an IgM binding rosetting P. falciparum line TM284R+, which is a culture-adapted parasite derived from a Thai patient with cerebral malaria [12]. Rosetting is the binding of iRBC to two or more uninfected RBC, and is a PfEMP1-mediated parasite virulence phenotype that is implicated in severe malaria [13]. Many rosetting PfEMP1 variants bind IgM [14], and the IgM is thought to strengthen and stabilise the rosettes [12,15]. We identified the PfEMP1 variant expressed by IgM binding rosetting TM284R+ parasites as TM284var1, and showed that the IgM binding region is the fourth DBL domain from the N-terminus, DBL4ζ [2] (Fig. 1A). This domain was initially described as a DBLβ subtype, however, more recent analyses indicate that this domain is a DBLζ subtype [6]. Henceforth, we shall refer to this domain as TM284var1 DBL4ζ.


Identification of the minimal binding region of a Plasmodium falciparum IgM binding PfEMP1 domain.

Semblat JP, Ghumra A, Czajkowsky DM, Wallis R, Mitchell DA, Raza A, Rowe JA - Mol. Biochem. Parasitol. (2015)

Identification of the minimal IgM binding region of the TM284var1 DBL4ζ domain.(A) Diagram showing the domain composition of the TM284var1 PfEMP1 variant, with the IgM binding DBL4ζ domain underlined in red. (B) Diagram showing the amino acid domain boundaries and IgM binding properties of each DBL4ζ deletion construct. The full length DBL4ζ domain (top bar) contains 16 cysteines as visualized by the dashed lines (construct used in previous work [2]). Seven deletion constructs were made spanning various regions of DBL4ζ. Proteins that bind human IgM are shown as black bars, and non-binding proteins are shown as white bars. (C) The minimal binding region of DBL4ζ (red). Subdomain (SD) 1 (yellow), 2 (blue) and 3 (grey) are shown, and cysteine residues are highlighted by arrowheads. The five charged residues within subdomain 2 predicted to be involved in IgM binding are shown in bold.
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fig0005: Identification of the minimal IgM binding region of the TM284var1 DBL4ζ domain.(A) Diagram showing the domain composition of the TM284var1 PfEMP1 variant, with the IgM binding DBL4ζ domain underlined in red. (B) Diagram showing the amino acid domain boundaries and IgM binding properties of each DBL4ζ deletion construct. The full length DBL4ζ domain (top bar) contains 16 cysteines as visualized by the dashed lines (construct used in previous work [2]). Seven deletion constructs were made spanning various regions of DBL4ζ. Proteins that bind human IgM are shown as black bars, and non-binding proteins are shown as white bars. (C) The minimal binding region of DBL4ζ (red). Subdomain (SD) 1 (yellow), 2 (blue) and 3 (grey) are shown, and cysteine residues are highlighted by arrowheads. The five charged residues within subdomain 2 predicted to be involved in IgM binding are shown in bold.
Mentions: Previously we studied an IgM binding rosetting P. falciparum line TM284R+, which is a culture-adapted parasite derived from a Thai patient with cerebral malaria [12]. Rosetting is the binding of iRBC to two or more uninfected RBC, and is a PfEMP1-mediated parasite virulence phenotype that is implicated in severe malaria [13]. Many rosetting PfEMP1 variants bind IgM [14], and the IgM is thought to strengthen and stabilise the rosettes [12,15]. We identified the PfEMP1 variant expressed by IgM binding rosetting TM284R+ parasites as TM284var1, and showed that the IgM binding region is the fourth DBL domain from the N-terminus, DBL4ζ [2] (Fig. 1A). This domain was initially described as a DBLβ subtype, however, more recent analyses indicate that this domain is a DBLζ subtype [6]. Henceforth, we shall refer to this domain as TM284var1 DBL4ζ.

Bottom Line: Here we studied an IgM binding domain from a rosette-mediating PfEMP1 variant, DBL4ζ of TM284var1, and found that the minimal IgM binding region mapped to the central region of the DBL domain, comprising all of subdomain 2 and adjoining parts of subdomains 1 and 3.Site-directed mutagenesis of charged amino acids within subdomain 2, predicted by molecular modelling to form the IgM binding site, showed no marked effect on IgM binding properties.Further work is needed to identify the specific interaction site for IgM within the minimal binding region of PfEMP1.

View Article: PubMed Central - PubMed

Affiliation: Institute of Immunology and Infection Research, Centre for Immunity, Infection and Evolution, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3FL, United Kingdom.

No MeSH data available.


Related in: MedlinePlus