S-layer, surface-accessible, and concanavalin A binding proteins of Methanosarcina acetivorans and Methanosarcina mazei

J Proteome Res. 2009 Apr;8(4):1972-82. doi: 10.1021/pr800923e.


The outermost cell envelope structure of many archaea and bacteria contains a proteinaceous lattice termed the surface layer or S-layer. It is typically composed of only one or two abundant, often posttranslationally modified proteins that self-assemble to form the highly organized arrays. Surprisingly, over 100 proteins were annotated to be S-layer components in the archaeal species Methanosarcina acetivorans C2A and Methanosarcina mazei Gö1, reflecting limitations of current predictions. An in vivo biotinylation methodology was devised to affinity tag surface-exposed proteins while overcoming unique challenges in working with these fragile organisms. Cells were adapted to growth under N2 fixing conditions, thus, minimizing free amines reactive to the NHS-label, and high pH media compatible with the acylation chemistry was used. A 3-phase separation procedure was employed to isolate intact, labeled cells from lysed-cell derived proteins. Streptavidin affinity enrichment followed by stringent wash conditions removed nonspecifically bound proteins. This methodology revealed S-layer proteins in M. acetivorans C2A and M. mazei Gö1 to be MA0829 and MM1976, respectively. Each was demonstrated to exist as multiple glycosylated forms using SDS-PAGE coupled with glycoprotein-specific staining, and by interaction with the lectin, Concanavalin A. A number of additional surface-exposed proteins and glycoproteins were identified and included all three subunits of the thermosome: the latter suggests that the chaperonin complex is both surface- and cytoplasmically localized. This approach provides an alternative strategy to study surface proteins in the archaea.

Publication types

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

MeSH terms

  • Archaeal Proteins / metabolism*
  • Chromatography, High Pressure Liquid
  • Concanavalin A / metabolism*
  • Glycoproteins / metabolism
  • Membrane Glycoproteins / metabolism*
  • Methanosarcina / metabolism*
  • Protein Binding
  • Proteome
  • Receptors, Concanavalin A / metabolism*
  • Tandem Mass Spectrometry


  • Archaeal Proteins
  • Glycoproteins
  • Membrane Glycoproteins
  • Proteome
  • Receptors, Concanavalin A
  • S-layer proteins
  • Concanavalin A