Detergent-assisted sample preparation for MALDI-MS: Investigation of octylglucoside and docecylmaltoside for matrix crystallization, on-plate digestion, and trypsin activity.

We show an easy and fast method for improved detection of lipophilic peptides with MALDI-MS utilizing the nonionic detergents n-octylglucoside and n-dodecylmaltoside (laurylmaltoside). Investigations comprised on-plate digestion of proteins with trypsin, detergent effects on the protease trypsin, and the changes in MALDI matrix crystallization. Investigations also exhibited a higher tryptic activity in trypsin activity assay of 139% when using laurylmaltoside as supplement. Crystallization changed toward a more homogeneous crystal distribution and especially trypsinized insulin spectra recorded with MALDI-MS showed improved detectability of lipophilic peptides.

Acyl modification and binding of mitochondrial ACP to multiprotein complexes.

The mitochondrial acyl carrier protein (ACPM/NDUFAB1) is a central element of the mitochondrial fatty acid synthesis type II machinery. Originally ACPM was detected as a subunit of respiratory complex I but the reason for the association with the large enzyme complex remained elusive. Complex I from the aerobic yeast Yarrowia lipolytica comprises two different ACPMs, ACPM1 and ACPM2. They are anchored to the protein complex by LYR (leucine-tyrosine-arginine) motif containing protein (LYRM) subunits LYRM3 (NDUFB9) and LYRM6 (NDUFA6). The ACPM1-LYRM6 and ACPM2-LYRM3 modules are essential for complex I activity and assembly/stability, respectively. We show that in addition to the complex I bound fraction, ACPM1 is present as a free matrix protein and in complex with the soluble LYRM4(ISD11)/NFS1 complex implicated in Fe-S cluster biogenesis. We show that the presence of a long acyl chain bound to the phosphopantetheine cofactor is important for docking ACPMs to protein complexes and we propose that association of ACPMs and LYRMs is universally based on a new protein-protein interaction motif.