Membrane fatty acid and phospholipid adaptation in bacteria in response to environmental stresses has been explored most extensively in E. coli
, B. subtilis
, Bacillus stearothermophilus
, and Acholeplasma laidlawii
). Until recently, the phenomena had not been explored as extensively in S. mutans
, an organism that inhabits a low-pH environment, dental plaque. Our interests have been focused on elucidating whether changes in membrane fatty acid composition occur (33
) and how and when they may occur (19
). From these studies, it was observed that the membrane composition of S. mutans
is altered in response to acidification; this shift can occur through the self-generation of acid by the organism, and it can be blocked by the fatty acid biosynthesis inhibitor cerulenin (19
). However, the mechanism by which the changes occurred was unclear.
Using results from a recent, elegant study in which the biochemical activity of FabM in S. pneumoniae
was examined (24
), we were able to identify and disrupt the homologue in S. mutans
UA159. The resultant mutant strain was extremely acid sensitive, but the mechanism by which monounsaturated fatty acids protect against acid is not yet understood. It may be possible that unsaturated bonds serve as a sink for protons; this, in conjunction with the activity of the F1
ATPase, could serve to protect the cytoplasmic contents from damage during growth under low-pH conditions.
S. mutans must survive low-pH environments because it produces acid via its metabolism of carbohydrates. The inability of the UR117 strain to metabolize glucose at pH values as low as those observed for UA159 may prevent the organism from acid adapting as fully as the wild type; thus, this may render the mutant strain more sensitive to extreme acidification. The simple addition of monounsaturated fatty acids to the growth medium of UR117 increased its ability to carry out glycolysis at low pH levels, most likely due to the incorporation of the fatty acids into the membrane.
Differences in glucose-specific PTS activity were expected, as it has been shown that the presence of exogenously supplied fatty acids can alter the PTS activity of S. mutans
). Enzyme IIC components of PTS are in the membrane; thus, shifting membrane composition could alter protein interactions and, consequently, affect activity. Despite increased glucose-specific PTS activity in UR117, these cultures were unable to perform glycolysis at pH values as low as UA159. The decreased glycolytic ability of UR117 is likely due to other enzymes further downstream of glucose transport.
The enhanced ATPase activity observed in UR117 was unexpected. The activity of F1
ATPase was higher in cells grown under low-pH conditions, where the membrane composition would have larger amounts of monounsaturated fatty acids (5
). In addition, we and others have shown evidence of transcriptional regulation of F-ATPase in streptococci during growth at low pH (21
). It is possible that the effect of a narrower ΔpH in UR117 had led to disregulation of the F-ATPase operon. We are currently investigating this possibility.
Differences in the membrane fatty acid composition may have additional effects on the phospholipid composition, peptidoglycan production, and other forms of membrane modifications. In addition, the effects of incorporation of exogenous fatty acids (in nutritionally supplemented strains) on other membrane components and the impact of this on de novo fatty acid biosynthesis, as well as other metabolic activities, are not completely understood. The presence of a putative transcriptional regulator, Smu1592 (2
), immediately upstream of fabH
leads to speculation of possible fatty acid biosynthesis regulation, which may occur when the organism is grown in an environment rich in fatty acids. Thus, many questions regarding the regulation of fatty acid biosynthesis and how membrane composition alterations may affect other membrane constituents need to be explored further to completely understand our observations.
Currently, our database searches as well as those done previously (24
) have indicated that FabM homologues exist only in streptococcal and staphylococcal species, with the exception of a homologue found in the gram-negative bacterium Fusobacterium nucleatum
(data not shown). Several recent articles have suggested the development of antibiotics to target fatty acid biosynthesis (13
). Bacterial fatty acid biosynthesis is classified as type II, in that each reaction is carried out by a separate enzyme. The fatty acid biosynthesis of eukaryotes, however, is classified as type I, in that all the reactions are carried out by a large enzyme complex (18
). The differences between the two fatty acid biosynthesis systems might be exploited, in that new drugs could be designed to target FabM enzymes (i.e., bacterial) that would probably not affect the enzymes used in eukaryotic cells. Thus, there is potential use in developing therapies that target ACP-isomerases, such as FabM.