Gas vesicles are proteinaceous, gas-filled nanostructures produced by some 
bacteria and archaea. The hydrophobic major structural protein GvpA forms the 
ribbed gas vesicle wall. An in-silico 3D-model of GvpA of the predicted 
coil-α1-β1-β2-α2-coil structure is available and implies that the two β-chains 
constitute the hydrophobic interior surface of the gas vesicle wall. To test the 
importance of individual amino acids in GvpA we performed 85 single 
substitutions and analyzed these variants in Haloferax volcanii ΔA + Amut 
transformants for their ability to form gas vesicles (Vac+ phenotype). In most 
cases, an alanine substitution of a non-polar residue did not abolish gas 
vesicle formation, but the replacement of single non-polar by charged residues 
in β1 or β2 resulted in Vac- transformants. A replacement of residues near the 
β-turn altered the spindle-shape to a cylindrical morphology of the gas 
vesicles. Vac- transformants were also obtained with alanine substitutions of 
charged residues of helix α1 suggesting that these amino acids form salt-bridges 
with another GvpA monomer. In helix α2, only the alanine substitution of His53 
or Tyr54, led to Vac- transformants, whereas most other substitutions had no 
effect. We discuss our results in respect to the GvpA structure and data 
available from solid-state NMR.