Characterizing heterogeneous bacterial surface functional groups using discrete affinity spectra for proton binding

Here we report results from a quantitative investigation of the types and d ensities of proton binding sites on a bacterial surface, Bacillus subtilis, from replicate acid-base titrations on bacteria at two ionic strengths (0. 025 and 0.1 M). In contrast to the surface complexation modeling (SCM) appr oach developed and widely used for mineral, e.g., iron oxides, and more rec ently bacterial surfaces; we fit the data using the linear programming meth od (LPM). Our results using LPM indicate five discrete binding sites occurr ing on the surface of B, subtilis likely corresponding to carboxylic sites at low pK(a) values, phosphoric sites at near-neutral pK(a) values, and ami ne sites at high pK(a) values. Replicate titrations on subsamples from the same bacterial population indicated less variability than has been suggeste d for bacterial surfaces. Both the pK(a) and site density values were found to be dependent on ionic strength. Comparing the pK(a) values determined h ere with LPM for B. subtilis to those determined independently by using a f ixed three site SCM model shows excellent agreement with the common sites l ikely corresponding to carboxylic, phosphoryl, and amine groups. However, t he LPM approach identifies a further two sites as compared to the SCM appro ach. These results have an important implication. Surfaces of a given bacte rial strain have a quantifiable, characteristic geochemical reactivity refl ecting discrete sites that can be traced back in terms of function to the u nderlying, cell wall structure, a well-characterized phenomenon for most ba cteria. However, an important caveat of our findings is that the absolute d ensities of these sites are highly dependent on a suite of both microbiolog ical and system chemical parameters.