Structural and spectroscopic characterisation of a heme peroxidase from sorghum

A cationic class III peroxidase from Sorghum bicolor was purified to homogeneity. The enzyme contains a high-spin heme, as evidenced by UV-visible spectroscopy and EPR. Steady state oxidation of guaiacol was demonstrated and the enzyme was shown to have higher activity in the presence of calcium ions. A Fe(III)/Fe(II) reduction potential of -266 mV vs NHE was determined. Stopped-flow experiments with H2O2 showed formation of a typical peroxidase Compound I species, which converts to Compound II in the presence of calcium. A crystal structure of the enzyme is reported, the first for a sorghum peroxidase. The structure reveals an active site that is analogous to those for other class I heme peroxidase, and a substrate binding site (assigned as arising from binding of indole-3-acetic acid) at the γ-heme edge. Metal binding sites are observed in the structure on the distal (assigned as a Na(+) ion) and proximal (assigned as a Ca(2+)) sides of the heme, which is consistent with the Ca(2+)-dependence of the steady state and pre-steady state kinetics. It is probably the case that the structural integrity (and, thus, the catalytic activity) of the sorghum enzyme is dependent on metal ion incorporation at these positions

Screening of malting sorghum samples for lactic acid bacteria with potentials for antimicrobial activity

Three varieties of sorghum grains (SK 5912, KSV 400 and KSV 8) undergoing malting process were screened for lactic acid bacteria (LAB) with antimicrobial activity and characterized using standard methods. Out of fifty-seven lactic acid bacteria isolates, eighteen isolates with antimicrobial producing potential were selected for further study. The isolates displayed significant (p < 0.05) inhibitory activity against two indicator strains Escherichia coli ATCC 111755 and Staphylococcus aureus ATCC 12600. Eleven of these inhibitor-producing isolates secreted inhibitory compounds that were sensitive to catalase while compounds from the other seven isolates continued to display inhibitory effect against the indicator strains after treatment with catalase. The proteinaceous nature and inactivation by catalase of these inhibitory compounds from the seven bacteria identified them as bacteriocins. Based on standard biochemical and microbiological tests, the isolates were tentatively identified as belonging to Lactococcus spp., Leuconostoc spp., Lactobacillus spp. and Streptococcus spp. However, three isolates (GS3A, S6A and S10B) were tentatively identified as Lactobacillus reuteri, Lactobacillus fermentum and Lactobacillus acidophilus, respectively. LAB isolated from three varieties of sorghum grains undergoing malting exhibited the ability to produce bacteriocin and hydrogen peroxide