Smartphone goniometer has excellent reliability between novice and experienced physical therapists in assessing knee range of motion.

INTRODUCTION: Smartphone app-based goniometer (SG) are emerging as an alternative to Universal Goniometers (UG) in assessing joint range of motion (ROM). This study examined whether the experience level of examiner affected the reliability of assessing knee flexion (KF) and knee extension (KE) ROM using UG and SG. METHODS: Participants with osteoarthritis of the knee or following total knee replacement were recruited. KF and KE ROM using UG and SG were assessed twice by an experienced physical therapist (PT) and a student PT (SPT). Intraclass correlation coefficients (ICC) examined the interrater (experienced PT vs SPT) and intrarater reliabilities (for experienced PT and SPT) in assessing KF and KE ROM for UG and SG. Concurrent relationships were examined between the knee ROM with pain and physical function using Pearson Correlation Coefficient (r). RESULTS: The interrater reliability in assessing KF and KE ROM was excellent (ICC>0.90) between novice and experienced examiners. The standard error of measurement (SEM) for novice examiner in assessing KF was 1° and 2° while using UG and SG respectively; whereas the SEM for experienced examiner in assessing KF was 1° irrespective of which device was used. The concurrent relationships between KF and KE ROM with measures of pain and function were divergent (moderate to low correlations; r <0.7; p > 0.05). CONCLUSION: Both UG and SG have smaller measurement error in assessing KF and KE ROM irrespective of experience level of examiner and therefore no one tool is superior than the other for assessing knee ROM in clinical practice.

Free RNA polymerase in Escherichia coli.

The frequencies of transcription initiation of regulated and constitutive genes depend on the concentration of free RNA polymerase holoenzyme [Rf] near their promoters. Although RNA polymerase is largely confined to the nucleoid, it is difficult to determine absolute concentrations of [Rf] at particular locations within the nucleoid structure. However, relative concentrations of free RNA polymerase at different growth rates, [Rf]rel, can be estimated from the activities of constitutive promoters. Previous studies indicated that the rrnB P2 promoter is constitutive and that [Rf]rel in the vicinity of rrnB P2 increases with increasing growth rate. Recently it has become possible to directly visualize Rf in growing Escherichia coli cells. Here we examine some of the important issues relating to gene expression based on these new observations. We conclude that: (i) At a growth rate of 2 doublings/h, there are about 1000 free and 2350 non-specifically DNA-bound RNA polymerase molecules per average cell (12 and 28%, respectively, of 8400 total) which are in rapid equilibrium. (ii) The reversibility of the non-specific binding generates more than 1000 free RNA polymerase molecules every second in the immediate vicinity of the DNA. Of these, most rebind non-specifically to the DNA within a few ms; the frequency of non-specific binding is at least two orders of magnitude greater than specific binding and transcript initiation. (iii) At a given amount of RNA polymerase per cell, [Rf] and the density of non-specifically DNA-bound RNA polymerase molecules along the DNA both vary reciprocally with the amount of DNA in the cell. (iv) At 2 doublings/h an E. coli cell contains, on the average, about 1 non-specifically bound RNA polymerase per 9 kbp of DNA and 1 free RNA polymerase per 20 kbp of DNA. However some DNA regions (i.e. near active rRNA operons) may have significantly higher than average [Rf].

Medium-dependent control of the bacterial growth rate.

By combining results from previous studies of nutritional up-shifts we here re-investigate how bacteria adapt to different nutritional environments by adjusting their macromolecular composition for optimal growth. We demonstrate that, in contrast to a commonly held view the macromolecular composition of bacteria does not depend on the growth rate as an independent variable, but on three factors: (i) the genetic background (i.e. the strain used), (ii) the physiological history of the bacteria used for inoculation of a given growth medium, and (iii) the kind of nutrients in the growth medium. These factors determine the ribosome concentration and the average rate of protein synthesis per ribosome, and thus the growth rate. Immediately after a nutritional up-shift, the average number of ribosomes in the bacterial population increases exponentially with time at a rate which eventually is attained as the final post-shift growth rate of all cell components. After a nutritional up-shift from one minimal medium to another minimal medium of higher nutritional quality, ribosome and RNA polymerase syntheses are co-regulated and immediately increase by the same factor equal to the increase in the final growth rate. However, after an up-shift from a minimal medium to a medium containing all 20 amino acids, RNA polymerase and ribosome syntheses are no longer coregulated; a smaller rate of synthesis of RNA polymerase is compensated by a gradual increase in the fraction of free RNA polymerase, possibly due to a gradual saturation of mRNA promoters. We have also analyzed data from a recent publication, in which it was concluded that the macromolecular composition in terms of RNA/protein and RNA/DNA ratios is solely determined by the effector molecule ppGpp. Our analysis indicates that this is true only in special cases and that, in general, medium adaptation also depends on factors other than ppGpp.

Modulation of Chemical Composition and Other Parameters of the Cell at Different Exponential Growth Rates.

This review begins by briefly presenting the history of research on the chemical composition and other parameters of cells of E. coli and S. enterica at different exponential growth rates. Studies have allowed us to determine the in vivo strength of promoters and have allowed us to distinguish between factor-dependent transcriptional control of the promoter and changes in promoter activity due to changes in the concentration of free functional RNA polymerase associated with different growth conditions. The total, or bulk, amounts of RNA and protein are linked to the growth rate, because most bacterial RNA is ribosomal RNA (rRNA). Since ribosomes are required for protein synthesis, their number and their rate of function determine the rate of protein synthesis and cytoplasmic mass accumulation. Many mRNAs made in the presence of amino acids have strong ribosome binding sites whose presence reduces the expression of all other active genes. This implies that there can be profound differences in the spectrum of gene activities in cultures grown in different media that produce the same growth rate. Five classes of growth-related parameters that are generally useful in describing or establishing the macromolecular composition of bacterial cultures are described in detail in this review. A number of equations have been reported that describe the macromolecular composition of an average cell in an exponential culture as a function of the culture doubling time and five additional parameters: the C- and D-periods, protein per origin (PO), ribosome activity, and peptide chain elongation rate.

Feedback control of ribosome synthesis in Escherichia coli is dependent on eight critical amino acids.

When bacteria growing in minimal medium are supplied with exogenous amino acids, they respond by increasing the synthesis of ribosomes; this leads to more protein synthesis capacity and faster growth. To examine how amino acids control the synthesis of ribosomes, two strategies were used. First, single amino acids were added to bacteria growing in minimal medium and their effect on the relative strength of the rrnB P1 promoter was determined. The addition of any one of eight amino acids (alanine, glutamine, and glutamic acid, isoleucine, leucine, methionine, serine, valine) increased the strength of the P1 promoter by 1.25- to 2.0-fold with no appreciable effect on transcription from an isolated rrn P2 promoter or on the bacterial growth rate. The effects of adding combinations of these critical amino acids were partially additive. When any one of the other amino acids was added, no discernable stimulation in relative P1 expression or growth was observed. In the second strategy, all amino acids were present in the growth medium, but the carbon source was altered to change the growth rate. In this case the relative strength of the P1 promoter was always constant and maximal. We suggest that addition of any of the eight critical amino acids reduces the ppGpp synthesis activity of the spoT gene product; the lower ppGpp levels, in turn, increase the strength of the rrn P1 promoters. It is suggested that these amino acids are involved in a feedback chain of reactions that control the rate of ribosome function by adjusting the rate of ribosome synthesis.

Control of rRNA synthesis in Escherichia coli: a systems biology approach.

The first part of this review contains an overview of the various contributions and models relating to the control of rRNA synthesis reported over the last 45 years. The second part describes a systems biology approach to identify the factors and effectors that control the interactions between RNA polymerase and rRNA (rrn) promoters of Escherichia coli bacteria during exponential growth in different media. This analysis is based on measurements of absolute rrn promoter activities as transcripts per minute per promoter in bacterial strains either deficient or proficient in the synthesis of the factor Fis and/or the effector ppGpp. These absolute promoter activities are evaluated in terms of rrn promoter strength (V(max)/K(m)) and free RNA polymerase concentrations. Three major conclusions emerge from this evaluation. First, the rrn promoters are not saturated with RNA polymerase. As a consequence, changes in the concentration of free RNA polymerase contribute to changes in rrn promoter activities. Second, rrn P2 promoter strength is not specifically regulated during exponential growth at different rates; its activity changes only when the concentration of free RNA polymerase changes. Third, the effector ppGpp reduces the strength of the rrn P1 promoter both directly and indirectly by reducing synthesis of the stimulating factor Fis. This control of rrn P1 promoter strength forms part of a larger feedback loop that adjusts the synthesis of ribosomes to the availability of amino acids via amino acid-dependent control of ppGpp accumulation.