@article {5053, title = {Detection of noncovalent complex between alpha-amylase and its microbial inhibitor tendamistat by electrospray ionization mass spectrometry}, journal = {Rapid Communications in Mass Spectrometry}, volume = {15}, number = {2}, year = {2001}, note = {ISI Document Delivery No.: 396KYTimes Cited: 11Cited Reference Count: 58}, pages = {89-96}, type = {Article}, abstract = {Electrospray ionization mass spectrometry (ESI-MS) is now routinely used for detection of noncovalent complexes. However, detection of noncovalent protein-protein complexes is not a widespread practice and still produces some challenges for mass spectrometrists. Here we demonstrate the detection of a noncovalent protein-protein complex between alpha -amylase and its microbial inhibitor tendamistat using ESI-MS. Crude porcine pancreatic alpha -amylase was purified using a glycogen precipitation method. Noncovalent complexes between porcine pancreatic alpha -amylase and its microbial inhibitor tendamistat are probed and detected using ESI-MS. The atmosphere-vacuum ESI conditions along with solution conditions and the ratio of inhibitor over enzyme strongly affect the detection of noncovalent complexes in the gas phase. ESI mass spectra of alpha -amylase at pH 7 exhibited charge states significantly lower than that reported previously, which is indicative of a native protein conformation necessary to produce a noncovalent complex. Detection of noncovalent complexes in the gas phase suggests that further use of conventional biochemical approaches to provide a qualitative, and in some cases even quantitative, characterization of equilibria of noncovalent complexes in solution is possible. Copyright (C) 2001 John Wiley \& Sons, Ltd.}, keywords = {ANALYTICAL ULTRACENTRIFUGATION, BINDING, CRYSTAL-STRUCTURE, CYTOCHROME-C, GAS-PHASE, HEME, MYOGLOBIN, PROBING CONFORMATIONAL-CHANGES, PROTEIN PROTEINASE-INHIBITORS, RECOGNITION SITES}, isbn = {0951-4198}, url = {://000166636600003}, author = {Douglas, D. J. and Collings, B. A. and Numao, S. and Nesatyy, V. J.} }