Characterization of Anopheles gambiae transglutaminase 3 (AgTG3) and its native substrate Plugin

J Biol Chem. 2013 Feb 15;288(7):4844-53. doi: 10.1074/jbc.M112.435347. Epub 2013 Jan 3.

Abstract

Male Anopheles mosquitoes coagulate their seminal fluids via cross-linking of a substrate, called Plugin, by the seminal transglutaminase AgTG3. Formation of the "mating plug" by cross-linking Plugin is necessary for efficient sperm storage by females. AgTG3 has a similar degree of sequence identity (~30%) to both human Factor XIII (FXIII) and tissue transglutaminase 2 (hTG2). Here we report the solution structure and in vitro activity for the cross-linking reaction of AgTG3 and Plugin. AgTG3 is a dimer in solution and exhibits Ca(2+)-dependent nonproteolytic activation analogous to cytoplasmic FXIII. The C-terminal domain of Plugin is predominantly α-helical with extended tertiary structure and oligomerizes in solution. The specific activity of AgTG3 was measured as 4.25 × 10(-2) units mg(-1). AgTG3 is less active than hTG2 assayed using the general substrate TVQQEL but has 8-10× higher relative activity when Plugin is the substrate. Mass spectrometric analysis of cross-linked Plugin detects specific peptides including a predicted consensus motif for cross-linking by AgTG3. These results support the development of AgTG3 inhibitors as specific and effective chemosterilants for A. gambiae.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Anopheles / enzymology*
  • Calcium / chemistry
  • Cross-Linking Reagents / chemistry
  • Cytoplasm / metabolism
  • Dimerization
  • Female
  • Male
  • Mass Spectrometry / methods
  • Models, Chemical
  • Molecular Sequence Data
  • Peptides / chemistry
  • Protein Binding
  • Protein Glutamine gamma Glutamyltransferase 2
  • Protein Structure, Tertiary
  • Sequence Homology, Amino Acid
  • Structure-Activity Relationship
  • Transglutaminases / chemistry*
  • Transglutaminases / metabolism

Substances

  • Cross-Linking Reagents
  • Peptides
  • TGM2 protein, human
  • Protein Glutamine gamma Glutamyltransferase 2
  • Transglutaminases
  • Calcium