(Received for publication, May 14, 1996)

Koen M. G. M. van der Drift , Herman P. Spaink ^§ , Guido V. Bloemberg ^§ , Anton A. N. van Brussel ^§ , Ben J. J. Lugtenberg ^§ , Johan Haverkamp and Jane E. Thomas-Oates

From the  Department of Mass Spectrometry, Utrecht University, F.A.F.C. Went Gebouw, Sorbonnelaan 16, 3584 CA Utrecht, The Netherlands and the ^§ Institute of Molecular Plant Sciences, Clusius Laboratory, Leiden University, Wassenaarseweg 64, 2333 AL Leiden, The Netherlands

The lipo-chitin oligosaccharides (LCO) or nodulation factors synthesized by Rhizobium leguminosarum bv. trifolii were analyzed using positive mode fast atom bombardment and positive and negative mode electrospray ionization mass spectrometry. From their mass spectrometric behavior it is clearly possible to distinguish between the [M + Na]⁺ pseudomolecular ion of the nodE-independent molecule IV(C18:1,Ac) and the [M + H]⁺ pseudomolecular ion of the nodE-dependent molecule IV(C20:4,Ac), although they both have the same mass value. The results unequivocally show that the bacterial strain investigated produces nodE-dependent LCOs with highly unsaturated fatty acyl moieties. We further demonstrate that the interpretation of the mass spectrometric data by Philip-Hollingsworth et al. (Philip-Hollingsworth, S., Orgambide, G. G., Bradford, J. J., Smith, D. K., Hollingsworth, R. I., and Dazzo, F. B. (1995) J. Biol. Chem. 270, 20968) is incorrect and that their data do not contradict our hypothesis that the nodE gene determines the host specificity of R. leguminosarum bv. trifolii.

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