Seasonal patterns of CO2 and water vapor exchange of three salt marsh succulents

Summary Diurnal carbon dioxide exchange patterns of three salt marsh succulents, Borrichia frutescens, Batis maritima and Salicornia virginica, were determined on a seasonal basis in the marsh at Sapelo Island, Georgia. Year-round photosynthetic activity was observed in these species though winter rates of CO₂ exchange were reduced. Net primary productivity, estimated using gas exchange techniques, agreed with previously reported harvest data. The role of light and temperature in the control of seasonal photosynthetic changes was investigated. A similar variation in light utilization with season was found in all three species, while seasonal temperature acclimation was species dependent. Less than 20% of fixed CO₂ was lost through dark respiration in any of the species.Water use in the salt marsh succulents was found to be relatively inefficient. High rates of transpiration were observed both summer and winter in the succulents.The succulents were judged to be C₃ plants on the basis of several criteria.Contribution No. 391 from the University of Georgia Marine Institute     

Mutants of Cellulomonas which produce increased levels of β-glucosidase

Summary Mutants have been isolated from a strain of Cellulomonas which are capable of producing up to 26-fold higher levels of -glucosidase than the parent under certain growth conditions.     

Apparent fusion of the TOL plasmid with the R91 drug resistance plasmid in Pseudomonas aeruginosa.

The TOL catabolic plasmid was shown to be compatible with the R91 drug resistance plasmid. However, the TOL plasmid was extremely unstable in mutant PA03 of P. aeruginosa. By selecting for stabilization of the TOL plasmid in PA03 harbouring R91, it was possible to isolate a strain in which markers from both R91 and TOL appeared to exist in a single recombinant plasmid. This plasmid, pND3, encoded resistance to carbenicillin, was able to transfer at the same frequency as the R91 plasmid and encoded the ability to grow on m-toluate, p-toluate, m-xylene, p-xylene and toluene. In addition, it was shown to be incompatible with the NAH catabolic plasmid and it could be transferred by transduction. The TOL plasmid could stabilize in PA03 harbouring R91 without fusion with R91, and could stabilize in PA03 in the absence of R91. PA03 harbouring either the recombinant plasmid or the stable TOL plasmid in the absence of R91 could promote bacterial chromosome transfer between mutant derivatives of P. aeruginosa strain PA0.     

The two-step interaction between α-dimethylaminonaphthalene-1-sulfonyl-pepsinogen-(1–12) and pepsin

The binding between α-dimethylaminonaphthalenesulfonyl-(1–12) and porcine pepsin can be detected by the large changes that occur in the fluorescence spectra of the dimethylaminonaphthalenesulfonyl chromophore due to energy transfer from tryptophan residues of the protein. The interaction was previously shown to consist of two steps: a fast step leading to a greatly enhanced fluorescence followed by a slower rearrangement step which reduces the fluorescence but leads to tighter binding and inhibition of the catalytic activity of pepsin (1). The two steps have been studied over a wide range of values of pH, temperature, and ionic strength to gain additional insights into the physical events occurring during the interaction. Based on the pH and ionic strength dependence, the initial step most likely involves electrostatic interaction of the basic peptide inhibitor with the acidic surface of pepsin in a rapid collision process. The use of this fluorescent reporter group has also suggested that the equilibrium binding after the slower rearrangement may also be pH dependent with most effective binding at higher pH. The kinetics of the slow step were measured by monitoring the continuous fluorescence decay. The resulting rates are compared to the rates observed by others for binding of pepstatin to pepsin. From the pH dependence of fluorescence, pK_app values are obtained for the dansylated peptide (3.25), for the pH dependence of the initial binding step (4.87), and for the equilibrium position (4.75).     

Kinetics of biofilm nitrification

The reaction rates (r(NH(4) (+) ) and r(NO(2) (-) )) in the two-step nitrification reaction were measured in a fluidized-sand-bed biofilm reactor under a range of steady-state conditions with respect to bulk NH(4) (+), NO(2) (-), and O(2) concentrations. It was shown from theory and experiment that under low NH(4) (+) concentration conditions, if the O(2)/NH(4) (+) concentration ratio in the bulk liquid is less than the stoichiometric coefficient (3.4 mg/mg), then oxygen will be rate limiting. In all experiments r(NO(2) (-) ) decreased more than r(NH(4) (+) ) under low oxygen conditions. This resulted in high NO(2) (-) effluent concentrations under low residence time conditions. The influence of the oxygen penetration effects on the relative values of r(NH(4) (+) ) and r(NO(2) (-) ) was experimentally shown to be caused either by the Nitrobacter location in the inner biofilm regions or by a K(m) effect for oxygen. Theoretical support of these findings was provided by a differential diffusion-reaction model which was used to simulate the experimental results.     

Replacement of lysine 269 by arginine in Escherichia coli tryptophan indole-lyase affects the formation and breakdown of quinonoid complexes

Lysine 269 in Escherichia coli tryptophan indole-lyase (tryptophanase) has been changed to arginine by site-directed mutagenesis. The resultant K269R mutant enzyme exhibits kcat values about 10% those of the wild-type enzyme with S-(o-nitrophenyl)-L-cysteine, L-tryptophan, and S-benzyl-L-cysteine, while kcat/Km values are reduced to 2% or less. The pH profile of kcat/Km for S-benzyl-L-cysteine for the mutant enzyme exhibits two pK alpha values which are too close to separate, with an average value of 7.6, while the wild-type enzyme exhibits pK alpha values of 6.0 and 7.8. The pK alpha for the interconversion of the 335 and 412 nm forms of the K269R enzyme is 8.3, while the wild-type enzyme exhibits a pK alpha of 7.4. Steady-state kinetic isotope effects on the reaction of [alpha-2H]S-benzyl-L-cysteine with the K269R mutant enzyme (Dkcat = 2.0; D(kcat/Km) = 3.9) are larger than those of the wild-type enzyme (Dkcat = 1.4; D(kcat/Km) = 2.9). Rapid scanning stopped-flow kinetic studies demonstrate that the K269R mutant enzyme does not accumulate quinonoid intermediates with L-alanine, L-tryptophan, or S-methyl-L-cysteine, but does form quinonoid absorption peaks in complexes with S-benzyl-L-cysteine and oxidolyl-L-alanine, whereas wild-type enzyme forms prominent quinonoid bands with all these amino acids. Single wavelength stopped-flow kinetic studies demonstrate that the alpha-deprotonation of S-benzyl-L-cysteine is 6-fold slower in the K269R mutant enzyme, while the intrinsic deuterium kinetic isotope effect is less for the K269R enzyme (Dk = 4.2) than for the wild-type (Dk = 7.9). The decay of the K269R quinonoid intermediate in the presence of benzimidazole is 7.1-fold slower than that of the wild-type enzyme. These results demonstrate that Lys-269 plays a significant role in the conformational changes or electrostatic effects obligatory to the formation and decomposition of the quinonoid intermediate, although it is not an essential basic residue.     

Spontaneous Mutation in the Escherichia Coli Laci Gene

To gain more detailed insight into the nature and mechanisms of spontaneous mutations, we undertook a DNA sequence analysis of a large collection of spontaneous mutations in the N-terminal region of the Escherichia coli lacI gene. This region of circa 210 base pairs is the target for dominant lacI mutations (i-d) and is suitable for studies of mutational specificity since it contains a relatively high density of detectable mutable sites. Among 414 independent i-d mutants, 70.8% were base substitutions, 17.2% deletions, 7.7% additions and 4.3% single-base frameshifts. The base substitutions were both transitions (60%) and transversions (40%), the largest single group being G.C----A.T (47% of base substitutions). All four transversions were observed. Among the 71 deletions, a hotspot (37 mutants) was present: an 87-bp deletion presumably directed by an 8-bp repeated sequence at its endpoints. The remaining 34 deletions were distributed among 29 different mutations, either flanked (13/34) or not flanked (21/34) by repeated sequences. The 32 additions comprised 29 different events, with only two containing a direct repeat at the endpoints. The single-base frameshifts were the loss of a single base from either repeated (67%) or nonrepeated (33%) bases. A comparison with the spectrum obtained previously in strains defective in DNA mismatch correction (mutH, mutL, mutS strains) yielded information about the apparent efficiency of mismatch repair. The overall effect was 260-fold but varied substantially among different classes of mutations. An interesting asymmetry was uncovered for the two types of transitions, A.T----G.C and G.C----A.T being reduced by mismatch repair 1340- and 190-fold, respectively. Explanations for this asymmetry and its possible implications for the origins of spontaneous mutations are discussed.