Saturating proteome identification and the study of post-translational protein modifications of Acholeplasma laidlawii using combination of single- and two-dimension gel electrophoresis followed by mass-spectrometry analysis have been carried out. Results were compared to the earlier identified proteome of Mycoplasma gallisepticum. It was found that M. gallisepticum and A. laidlawii express 61 and 58% of the annotated ORFs respectively. All subunits of DNA-polymerase III were identified during our study which indicates that our methods can detect single copies of a given protein per cell. Metabolic pathways of the respective mycoplasmas were compared further in this work.