In both prokaryotes and eukaryotes methyl groups can be added to and removed from the carboxyl groups of proteins. Recent work has revealed that these reactions have a role in several behavioural phenomena. The nature of this role has been uncovered in one case--that of bacterial chemotaxis.

The properties of two classes of behavioral mutants of Escherichia coli (called tsr and tar) The costs of publication of this article were defrayed in part by the payment of page charges from funds made available to support the research which is the subject of the article. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.grade. Nickel and cobalt were used as the sulfate salts; aspartate, acetate, and benzoate, as the sodium salts. Bacteria. Chemotactic mutants AW518 and AW539 were isolated from the chemotactically wild-type Escherschia coli K-12 strain AW405 as previously described (5, 6). The double mutant AW569 was constructed by transferring the chemotaxis defect of AW518 to AW539 via cotransduction with thr from a thr+ derivative of AW518. Behavioral Assays. Bacteria used in capillary assays (7) were grown on minimal medium and washed as described previously (7). Responses were quantitated by integrating the areas under concentration-response curves. Bacteria used in temporal assays (2,8,9) were grown on tryptone broth as described previously (10). Five milliliters of cells were washed three times in 10 mM potassium phosphate buffer (pH 7.0) containing 0.1 mM EDTA (chemotaxis medium) and resuspended in this medium. Temporal assays were quantitated by measuring the duration of the response to stimuli; the response to attractants is a suppression pf tumbling (8), while the response to repellents is an enhancement of tumbling (2).Methylation. Cells were grown and washed as described previously (4). All subsequent incubations were carried out at 300 with rotary shaking. Washed cells were resuspended (to 3 X 108 cells per ml) in chemotaxis medium containing chloramphenicol (200 tg/ml) to inhibit protein synthesis, sodium D,L-lactate (10 mM), and L-threonine, L-leucine, and L-histidine (0.1 mM each). After 5 min of incubation [3H]methionine was added (4 ,tM) and 5-ml aliquots were distributed to various flasks. Samples to be stimulated with attractants or repellents received these compounds 30 or 58 min later, respectively. All reactions were terminated after 60 min of labeling by the addition of 0.25% formalin (final concentration). These times of incubation (30 min for attractants and 2 min for repellents) allow the new equilibrium levels of methylation to be established before termination of the experiment. § Samples were centrifuged to sediment the cells and the pellets were resuspended in 100 ,l of Laemmli sample buffer (11)

The American lobster Homarus americanus is a decapod crustacean with both high economic and scientific importance. To facilitate physiological investigations of peptide transmitter/hormone function in this species, we have used matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS), matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and nanoscale liquid chromatography coupled to electrospray ionization quadrupole time-of-flight tandem mass spectrometry (nanoLC-ESI-Q-TOF MS/MS) to elucidate the peptidome present in its nervous system and neuroendocrine organs. In total, 84 peptides were identified, including 27 previously known H. americanus peptides (e.g., VYRKPPFNGSIFamide [Val(1)-SIFamide]), 23 peptides characterized previously from other decapods, but new to the American lobster (e.g., pQTFQYSRGWTNamide [Arg(7)-corazonin]), and 34 new peptides de novo sequenced/detected for the first time in this study. Of particular note are a novel B-type allatostatin (TNWNKFQGSWamide) and several novel FMRFamide-related peptides, including an unsulfated analog of sulfakinin (GGGEYDDYGHLRFamide), two myosuppressins (QDLDHVFLRFamide and pQDLDHVFLRFamide), and a collection of short neuropeptide F isoforms (e.g., DTSTPALRLRFamide and FEPSLRLRFamide). Our data also include the first detection of multiple tachykinin-related peptides in a non-brachyuran decapod, as well as the identification of potential individual-specific variants of orcokinin and orcomyotropin-related peptide. Taken collectively, our results not only expand greatly the number of known H. americanus neuropeptides, but also provide a framework for future studies on the physiological roles played by these molecules in this commercially and scientifically important species.

A protein methylation reaction involved in chemotaxis of Escherichia coli has been identified. The involvement of this reaction in chemotaxis in indicated by four lines of evidence. (a) The methylation reaction is altered in several classes of generally nonchemotactic mutants and is coreverted with the chemotaxis defects. (b) The methylation level of the protein is affected by chemotactic stimuli. (c) The transferred methyl group is derived from methionine and is labile, in accord with the known fact that chemotaxis requires a continuous supply of methionine. (d) Methylation is abnormal in various mutants having defective or missing flagella.