Contrary to the prevalent view, there seems to be no competition in the mammalian female genital tract among large numbers of sperm cells that are racing towards the egg. Instead, small numbers of the ejaculated sperm cells enter the Fallopian tube, and these few must be guided to make the remaining long, obstructed way to the egg. Here, we review the mechanisms by which mammalian sperm cells are guided to the egg.

Regulation of the direction of flagellar rotation is central to the mechanism of bacterial chemotaxis. The transitions between counterclockwise and clockwise rotation are controlled by a "switch complex" composed of three proteins (FUG, FHlM, and FUN) CheY (8,9). In this way, the flux of information from the receptors is integrated into a common form: phosphorylated CheY. Changes in the phosphorylation level ofCheY are believed to be sensed by a group ofproteins located on the cytoplasmic face of the flagellar motor known as the "switch complex" (10), which causes the motor to adopt either a clockwise bias (if levels of phosphorylated CheY are high) or a counterclockwise bias (when phosphorylated CheY is low) (11). The mechanism of switch function is not known. Overproduction of CheY in a strain lacking all other chemotaxis proteins (12-14) or insertion of purified CheY into cell envelopes devoid of cytoplasm (15) MATERIALS AND METHODSPreparation of CheY Beads. CheY beads and control beads were prepared in parallel under identical conditions. Two samples (1-g dry weight each) of CNBr-activated Sepharose 4B beads (Pharmacia) were washed five times in 14 ml of 1 mM HCl and then three times in coupling buffer (0.1 M NaHC03/0.5 M NaCl, pH 8.3). CheY [5-45 mg, determined from absorbance at 280 nm by using a molar extinction coefficient of 6970 M-1"cm-1 (19)] purified from Escherichia coli (11) was dialyzed twice against 2 liters of coupling buffer and added to one tube. An equal volume of coupling buffer was added to the other tube (containing control beads). Both tubes were mixed end-over-end overnight at 4°C. In this way 95-99% ofthe CheY became covalently immobilized. Bovine serum albumin (BSA, 20 mg), previously dialyzed against coupling buffer, was then added to both tubes in order to block unreacted cyano groups, and the incubation with mixing continued for 5-10 hr at 4°C. To complete the coupling, additional BSA (20 mg), this time in 0.1 M Tris HCl (pH 7.9), was added to both tubes and incubated further for 5-10 hr. Finally the beads were washed in 50 mM Tris-HCl (pH 7.9) and stored at 4°C. Prior to use in the binding assay, the beads were washed and resuspended in 8-12 ml of Tris buffer. CheY57DE was produced and isolated as described earlier (20)

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Acetyl coenzyme A synthetase (Acs) activates acetate to acetyl coenzyme A through an acetyladenylate intermediate; two other enzymes, acetate kinase (Ack) and phosphotransacetylase (Pta), activate acetate through an acetyl phosphate intermediate. We subcloned acs, the Escherichia coli open reading frame purported to encode Acs (F. R. Blattner, V. Burland, G. Plunkett III, H. J. Sofia, and D. L. Daniels, Nucleic Acids Res. 21:5408-5417, 1993). We constructed a mutant allele, ⌬acs::Km, with the central 0.72-kb BclI-BclI portion of acs deleted, and recombined it into the chromosome. Whereas wild-type cells grew well on acetate across a wide range of concentrations (2.5 to 50 mM), those deleted for acs grew poorly on low concentrations (Յ10 mM), those deleted for ackA and pta (which encode Ack and Pta, respectively) grew poorly on high concentrations (Ն25 mM), and those deleted for acs, ackA, and pta did not grow on acetate at any concentration tested. Expression of acs from a multicopy plasmid restored growth to cells deleted for all three genes. Relative to wild-type cells, those deleted for acs did not activate acetate as well, those deleted for ackA and pta displayed even less activity, and those deleted for all three genes did not activate acetate at any concentration tested. Induction of acs resulted in expression of a 72-kDa protein, as predicted by the reported sequence. This protein immunoreacted with antiserum raised against purified Acs isolated from an unrelated species, Methanothrix soehngenii. The purified E. coli Acs then was used to raise anti-E. coli Acs antiserum, which immunoreacted with a 72-kDa protein expressed by wild-type cells but not by those deleted for acs. When purified in the presence, but not in the absence, of coenzyme A, the E. coli enzyme activated acetate across a wide range of concentrations in a coenzyme A-dependent manner. On the basis of these and other observations, we conclude that this open reading frame encodes the acetate-activating enzyme, Acs.Escherichia coli cells activate acetate to acetyl coenzyme A (acetyl-CoA) by two distinct pathways (Fig. 1) Brown et al. (6) hypothesized that the Acs pathway functions as a catabolite-repressible, acetate-inducible, high-affinity acetate uptake system that scavenges acetate present extracellularly at relatively low concentrations. They also proposed that the Ack-Pta pathway functions primarily in a catabolic role, excreting acetate and generating ATP during mixed-acid fermentation and aerobic growth on excess glucose or other glycolytic intermediates. Finally, they argued that the low-affinity Ack-Pta pathway activates acetate only when that molecule is present extracellularly in large quantity.In addition to their role in acetate metabolism, the acetate activation pathways have been implicated in the regulation of signal transduction by two-component regulatory systems in several bacterial species (reviewed in references 32 and 54; see also references 2, 9, 39, and 55), the regulation of the glucose starvation stimulon of E. coli ...

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