Image quality of iterative reconstruction in cranial CT imaging: comparison of model-based iterative reconstruction (MBIR) and adaptive statistical iterative reconstruction (ASiR).

OBJECTIVES: The purpose of this study was to compare cranial CT (CCT) image quality (IQ) of the MBIR algorithm with standard iterative reconstruction (ASiR). METHODS: In this institutional review board (IRB)-approved study, raw data sets of 100 unenhanced CCT examinations (120 kV, 50-260 mAs, 20 mm collimation, 0.984 pitch) were reconstructed with both ASiR and MBIR. Signal-to-noise (SNR) and contrast-to-noise (CNR) were calculated from attenuation values measured in caudate nucleus, frontal white matter, anterior ventricle horn, fourth ventricle, and pons. Two radiologists, who were blinded to the reconstruction algorithms, evaluated anonymized multiplanar reformations of 2.5 mm with respect to depiction of different parenchymal structures and impact of artefacts on IQ with a five-point scale (0: unacceptable, 1: less than average, 2: average, 3: above average, 4: excellent). RESULTS: MBIR decreased artefacts more effectively than ASiR (p < 0.01). The median depiction score for MBIR was 3, whereas the median value for ASiR was 2 (p < 0.01). SNR and CNR were significantly higher in MBIR than ASiR (p < 0.01). CONCLUSIONS: MBIR showed significant improvement of IQ parameters compared to ASiR. As CCT is an examination that is frequently required, the use of MBIR may allow for substantial reduction of radiation exposure caused by medical diagnostics. KEY POINTS: • Model-Based iterative reconstruction (MBIR) effectively decreased artefacts in cranial CT. • MBIR reconstructed images were rated with significantly higher scores for image quality. • Model-Based iterative reconstruction may allow reduced-dose diagnostic examination protocols.

High resolution compression elastography and color doppler sonography in characterization of breast fibroadenoma.

PURPOSE: To evaluate the use of color Doppler sonography and free hand sonoelastography in the assessment of breast fibroadenomas according to their size and shape. MATERIALS AND METHODS: From December 2012 to March 2013 women with 16 solid breast masses, classified as BI-RADS category 3 or 4 were examined with B-mode ultrasound, sonoelastography and color Doppler sonography. Lesions were subdivided according to their shape in round, ovoid or macrolobulated and according to their size (<2.0 cm or ≥2.0 cm). Two readers assessed sonoelastographic findings at 12.5 MHz using the tsukuba elasticity score and results of Doppler sonography using a score of 0, 1 or 2, depending on the degree of perfusion. RESULTS: Among the 16 examined lesions 3 showed a round shape, 9 were ovoid and in 4 cases a macrolobulated appearance could be seen. No significant differences concerning Tsukuba-score depending on various shapes of fibroadenomas in B-mode sonography could be shown (p = 0.91) and also comparison of Tsukuba-scores and size of masses revealed no significant differences (p = 1.0). Sizes of fibroadenomas ≥2 cm were significantly associated with an increased vascularization of the lesions (p = 0.04) and a macrolobulated appearance in B-mode sonography (p = 0.04). CONCLUSION: The combination of color Doppler sonography and sonoelastography in addition to B-mode sonography leads to an increased accuracy in distinguishing benign from malignant breast masses and to an improvement in characterization of fibroadenomas independent of their shape or size.

Upgrade to iterative image reconstruction (IR) in MDCT imaging: a clinical study for detailed parameter optimization beyond vendor recommendations using the adaptive statistical iterative reconstruction environment (ASIR) Part2: The chest.

PURPOSE: To compare the image quality in dose-reduced 64-row CT of the chest at different levels of adaptive statistical iterative reconstruction (ASIR) to full-dose baseline examinations reconstructed solely with filtered back projection (FBP) in a realistic upgrade scenario. MATERIALS AND METHODS: A waiver of consent was granted by the institutional review board (IRB). The noise index (NI) relates to the standard deviation of Hounsfield units in a water phantom. Baseline exams of the chest (NI = 29; LightSpeed VCT XT, GE Healthcare) were intra-individually compared to follow-up studies on a CT with ASIR after system upgrade (NI = 45; Discovery HD750, GE Healthcare), n = 46. Images were calculated in slice and volume mode with ASIR levels of 0 - 100 % in the standard and lung kernel. Three radiologists independently compared the image quality to the corresponding full-dose baseline examinations (-2: diagnostically inferior, -1: inferior, 0: equal, + 1: superior, + 2: diagnostically superior). Statistical analysis used Wilcoxon's test, Mann-Whitney U test and the intraclass correlation coefficient (ICC). RESULTS: The mean CTDIvol decreased by 53 % from the FBP baseline to 8.0 ± 2.3 mGy for ASIR follow-ups; p < 0.001. The ICC was 0.70. Regarding the standard kernel, the image quality in dose-reduced studies was comparable to the baseline at ASIR 70 % in volume mode (-0.07 ± 0.29, p = 0.29). Concerning the lung kernel, every ASIR level outperformed the baseline image quality (p < 0.001), with ASIR 30 % rated best (slice: 0.70 ± 0.6, volume: 0.74 ± 0.61). CONCLUSION: Vendors' recommendation of 50 % ASIR is fair. In detail, the ASIR 70 % in volume mode for the standard kernel and ASIR 30 % for the lung kernel performed best, allowing for a dose reduction of approximately 50 %.

Evaluation of a dedicated MDCT protocol using iterative image reconstruction after cervical spine trauma.

AIM: To evaluate radiation exposure for 64-row computed tomography (CT) of the cervical spine comparing two optimized protocols using filtered back projection (FBP) and adaptive statistical iterative reconstruction (ASIR), respectively. MATERIALS AND METHODS: Sixty-seven studies using FBP (scanner 1) were retrospectively compared with 80 studies using ASIR (scanner 2). The key scanning parameters were identical (120 kV dose modulation, 64 × 0.625 mm collimation, pitch 0.531:1). In protocol 2, the noise index (NI) was increased from 5 to 25, and ASIR and the high-definition (HD) mode were used. The scan length, CT dose index (CTDI), and dose-length product (DLP) were recorded. The image quality was analysed subjectively by using a three-point scale (0; 1; 2), and objectively by using a region of interest (ROI) analysis. Mann-Whitney U and Wilcoxon's test were used. RESULTS: In the FBP group, the mean CTDI was 21.43 mGy, mean scan length 186.3 mm, and mean DLP 441.15 mGy cm. In the ASIR group, the mean CTDI was 9.57 mGy, mean scan length 195.21 mm, and mean DLP 204.23 mGy cm. The differences were significant for CTDI and DLP (p < 0.001) and scan length (p = 0.01). There was no significant difference in the subjective image quality (p > 0.05). The estimated mean effective dose decreased from 2.38 mSv (FBP) to 1.10 mSv (ASIR). CONCLUSION: The radiation dose of 64-row MDCT can be reduced to a level comparable to plain radiography without loss of subjective image quality by implementation of ASIR in a dedicated cervical spine trauma protocol. These results might contribute to an improved relative risk-to-benefit ratio and support the justification of CT as a first-line imaging tool to evaluate cervical spine trauma.

[Evaluation of radiation dose in 64-row whole-body CT of multiple injured patients compared to 4-row CT]. / Evaluation der Strahlendosis bei Polytrauma-CT-Untersuchungen eines 64-Zeilen-CT im Vergleich zur 4-Zeilen-CT.

PURPOSE: To evaluate radiation exposure in whole-body CT (WBCT) of multiple injured patients comparing 4-row multidetector computed tomography (MDCT) to 64-row MDCT. MATERIALS AND METHODS: 200 WBCT studies were retrospectively evaluated: 92 4-row MDCT scans and 108 64-row MDCT scans. Each CT protocol was optimized for the particular CT system. The scan length, CT dose index (CTDI), and dose length product (DLP) were recorded and analyzed for radiation exposure. The mean effective dose was estimated based on conversion factors. Student's t-test was used for statistical analysis. RESULTS: The mean CTDIvol values (mGy) of the thorax and abdomen were significantly reduced with 64-row MDCT (10.2±2.5 vs. 11.4±1.4, p<0.001; 14.2±3.7 vs. 16.1±1.7, p<0.001). The DLP values (mGy×cm) of the head and thorax were significantly increased with 64-row MDCT (1305.9±201.1 vs. 849.8±90.9, p<0,001; 504.4±134.4 vs. 471.5±74.1, p=0.030). The scan lengths (mm) were significantly increased with 64-row MDCT: head 223.6±35.8 vs. 155.5±12.3 (p<0.001), thorax 427.4±44.5 vs. 388.3±57.5 (p<0.001), abdomen 520.3±50.2 vs. 490.8±51.6 (p<0.001). The estimated mean effective doses (mSv) were 22.4±2.6 (4-row MDCT) and 24.1±4.6 (64-row MDCT; p=0.001), resulting in a percentage increase of 8%. CONCLUSION: The radiation dose per slice of the thorax and abdomen can be significantly decreased by using 64-row MDCT. Due to the technical advances of modern 64-row MDCT systems, the scan field can be adapted to the clinical demands and, if necessary, enlarged without time loss. As a result, the estimated mean effective dose might be increased in WBCT.

Upgrade to iterative image reconstruction (IR) in abdominal MDCT imaging: a clinical study for detailed parameter optimization beyond vendor recommendations using the adaptive statistical iterative reconstruction environment (ASIR).

PURPOSE: To compare the image quality of dose-reduced 64-row abdominal CT reconstructed at different levels of adaptive statistical iterative reconstruction (ASIR) to full-dose baseline examinations reconstructed with filtered back-projection (FBP) in a clinical setting and upgrade situation. MATERIALS AND METHODS: Abdominal baseline examinations (noise index NI = 29; LightSpeed VCT XT, GE) were intra-individually compared to follow-up studies on a CT with an ASIR option (NI = 43; Discovery HD750, GE), n = 42. Standard-kernel images were calculated with ASIR blendings of 0 - 100 % in slice and volume mode, respectively. Three experienced radiologists compared the image quality of these 567 sets to their corresponding full-dose baseline examination (- 2: diagnostically inferior, - 1: inferior, 0: equal, + 1: superior, + 2: diagnostically superior). Furthermore, a phantom was scanned. Statistical analysis used the Wilcoxon - the Mann-Whitney U-test and the intra-class correlation (ICC). RESULTS: The mean CTDIvol decreased from 19.7 ±â€Š5.5 to 12.2 ±â€Š4.7 mGy (p < 0.001). The ICC was 0.861. The total image quality of the dose-reduced ASIR studies was comparable to the baseline at ASIR 50 % in slice (p = 0.18) and ASIR 50 - 100 % in volume mode (p > 0.10). Volume mode performed 73 % slower than slice mode (p < 0.01). CONCLUSION: After the system upgrade, the vendor recommendation of ASIR 50 % in slice mode allowed for a dose reduction of 38 % in abdominal CT with comparable image quality and time expenditure. However, there is still further dose reduction potential for more complex reconstruction settings.

[Angiographic diagnosis of acral circulatory disorders of the upper extremities]. / Angiographische Diagnostik akraler Durchblutungsstörungen an der oberen Extremität.

Acral ischemic lesions rarely affect the upper extremities. While in the lower limbs atherosclerosis is responsible for the majority of lesions, vasculitis and autoimmune diseases play an important role in the pathogenesis of ischemic lesions of the upper limbs. A considerable number of acral circulatory disorders present with Raynaud's phenomenon and often without associated necrosis. Raynaud's phenomenon is mainly idiopathic but may also be secondary to underlying conditions, such as autoimmune diseases and vasculitis. Because of its high spatial resolution and the often discrete morphological findings digital subtraction angiography (DSA) is still an important diagnostic method in the radiological evaluation of acral circulatory disorders of the hand. Angiographic features of vasculitis are not strictly pathognomonic but certain morphologic DSA findings are very typical and may allow for a radiologic diagnosis. For instance, atherosclerosis results in irregular contours of vessel walls in DSA in contrast to autoimmune diseases and vasculitis, which are usually characterized by smooth vessel walls and optional vasospasm, the latter being especially typical for thromboangiitis obliterans and scleroderma. In thromboangiitis obliterans occlusions of the distal hand arteries, corkscrew collateral vessels and subsequent development of fine collateral networks are typical findings. Abrupt or filiform occlusions of distal finger arteries with sparse collateralization and symmetric affection of both hands are suggestive of scleroderma. Disseminated segmental ectasis and stenosis as well as microaneurysms (63% of all patients) are very common in patients with panarteriitis nodosa.

Components of respiratory resistance monitored in mechanically ventilated patients.

The interrupter technique is commonly adopted to monitor respiratory resistance (Rrs,int) during mechanical ventilation; however, Rrs,int is often interpreted as an index of airway resistance (Raw). This study compared the values of Rrs,int provided by a Siemens 940 Lung Mechanics Monitor with total respiratory impedance (Zrs) parameters in 39 patients with normal spirometric parameters, who were undergoing elective coronary bypass surgery. Zrs was determined at the airway opening with pseudorandom oscillations of 0.2-6 Hz at end inspiration. Raw and tissue resistance (Rti) were derived from the Zrs data by model fitting; Rti and total resistance (Rrs,osc=Raw+Rti) were calculated at the actual respirator frequencies. Lower airway resistance (Rawl) was estimated by measuring tracheal pressure. Although good agreement was obtained between Rrs,osc and Rrs,int, with a ratio of 1.07+/-0.19 (mean+/-SD), they correlated poorly (r2=0.36). Rti and the equipment component of Raw accounted for most of Rrs,osc (39.8+/-11.9 and 43.0+/-6.9%, respectively), whereas only a small portion belonged to Rawl (17.2+/-6.3%). It is concluded that respiratory resistance may become very insensitive to changes in lower airway resistance and therefore, inappropriate for following alterations in airway tone during mechanical ventilation, especially in patients with relatively normal respiratory mechanics, where the tissue and equipment resistances represent the vast majority of the total resistance.

Comparative studies of the S0 and S2 multiline electron paramagnetic resonance signals from the manganese cluster in Photosystem II.

Electron paramagnetic resonance (EPR) spectroscopy is one of the major techniques used to analyse the structure and function of the water oxidising complex (WOC) in Photosystem II. The discovery of an EPR signal from the S0 state has opened the way for new experiments, aiming to characterise the S0 state and elucidate the differences between the different S states. We present a review of the biochemical and biophysical characterisation of the S0 state multiline signal that has evolved since its discovery, and compare these results to previous and recent data from the S2 multiline signal. We also present some new data from the S2 state reached on the second turnover of the enzyme.

Proton equilibria in the manganese cluster of photosystem II control the intensities of the S(0) and S(2) state g approximately 2 electron paramagnetic resonance signals.

We have studied the pH effect on the S(0) and S(2) multiline electron paramagnetic resonance (EPR) signals from the water-oxidizing complex of photosystem II. Around pH 6, the maximum signal intensities were detected. On both the acidic and alkaline sides of pH 6, the intensities of the EPR signals decreased. Two pKs were determined for the S(0) multiline signal; pK(1) = 4.2 +/- 0.2 and pK(2) = 8.0 +/- 0.1, and for the S(2) multiline signal the pKs were pK(1) = 4.5 +/- 0.1 and pK(2) = 7.6 +/- 0.1. The intensity of the S(0)-state EPR signal was partly restored when the pH was changed from acidic or alkaline pH back to pH approximately 6. In the S(2) state we observed partial recovery of the multiline signal when going from alkaline pH back to pH approximately 6, whereas no significant recovery of the S(2) multiline signal was observed when the pH was changed from acidic pH back to pH approximately 6. Several possible explanations for the intensity changes as a function of pH are discussed. Some are ruled out, such as disintegration of the Mn cluster or decay of the S states and formal Cl(-) and Ca(2+) depletion. The altered EPR signal intensities probably reflect the protonation/deprotonation of ligands to the Mn cluster or the oxo bridges between the Mn ions. Also, the possibility of decreased multiline signal intensities at alkaline pH as an effect of changed redox potential of Y(Z) is put forward.

Methanol modification of the electron paramagnetic resonance signals from the S(0) and S(2) states of the water-oxidizing complex of photosystem II

The Mn-derived electron paramagnetic resonance (EPR) multiline signal from the S(0) state of the water-oxidizing complex is observable only in the presence methanol. In the present study, we have characterized the effect of methanol on the EPR signals from the S(0) and S(2) states as well as on the EPR Signal II(slow) originating from the Tyrosine(D)(ox) radical. The amplitudes of the S(0) and S(2) multiline signals increase with the methanol concentration in a similar way, whereas the S(2) g=4.1 excited state signal amplitude shows a concomitant decrease. The methanol concentration at which half of the spectral change has occurred is approximately 0.2% and the effect is saturating around 5%. Methanol has an effect on the microwave power saturation of the S(2) multiline signal, as well. The microwave power at half saturation (P(1/2)) is 85 mW in the presence of methanol, whereas the signal relaxes much slower (P(1/2) approximately 27 mW) without. The relaxation of Signal II(slow) in the presence of methanol has also been investigated. The P(1/2) value of Signal II(slow) oscillates with the S cycle in a similar way as without methanol, but the P(1/2) values are consistently lower in the methanol-containing samples. From the results, we conclude that methanol modifies the magnetic properties of the S(0) and S(2) states in a similar way. The possible site and nature of methanol binding is discussed.

Comparison of the functional properties of the monomeric and dimeric forms of the isolated CP47-reaction center complex.

Chlorophyll fluorescence, thermoluminescence, and EPR spectroscopy have been used to investigate the functional properties of the monomeric and dimeric forms of the photosystem II CP47-reaction center (CP47-RC) subcore complex that was isolated (Zheleva, D., Sharma, J., Panico, M., Morris, H. R., and Barber, J. (1998) J. Biol. Chem. 273, 16122-16127). Chlorophyll fluorescence yield changes induced either by the initiation of continuous actinic light or by repetitive light flashes indicated that the dimeric, but not the monomeric, form of the CP47-RC complex showed secondary electron transport properties indicative of QA reduction. Thermoluminescence measurements also clearly distinguished the monomer from the dimer in that the latter showed a ZV band, which appeared at -55 degreesC, following illumination at -80 degreesC. This band has been determined to be an indicator of the photoaccumulation of QA-. The ability of the dimeric CP47-RC to show secondary electron transport properties was clearly demonstrated by EPR studies. The dimer was characterized by organic radical signals at about g = 2 induced either by illumination or by the addition of dithionite. The dithionite-induced signal was attributed to QA-, but there was no indication of any interaction with non-heme iron. The signal induced by light was more complex, being composed not only of the QA- radical but also of radicals generated on the donor side. Difference analyses indicated that one of these radicals is likely to be due to a D1 tyrosine 161 or D2 tyrosine 161. In contrast, the monomeric CP47-RC complex did not show similar EPR-detectable radicals and instead was dominated by a high yield of the spin-polarized triplet signal generated by recombination reactions between the oxidized primary reductant, pheophytin, and the primary donor, P680. It is also concluded from EPR analyses that both the monomeric and dimeric forms of the CP47-RC subcore complex contain one cytochrome b559 per reaction center. Overall the results suggest that photosystem II normally functions as a dimer complex and that monomerization at the level of the CP47-RC subcore complex leads to destabilization of the bound plastoquinone, which functions as QA.

Mutation of residue threonine-2 of the D2 polypeptide and its effect on photosystem II function in Chlamydomonas reinhardtii.

The D2 polypeptide of the photosystem II (PSII) complex in the green alga Chlamydomonas reinhardtii is thought to be reversibly phosphorylated. By analogy to higher plants, the phosphorylation site is likely to be at residue threonine-2 (Thr-2). We have investigated the role of D2 phosphorylation by constructing two mutants in which residue Thr-2 has been replaced by either alanine or serine. Both mutants grew photoautotrophically at wild-type rates, and noninvasive biophysical measurements, including the decay of chlorophyll fluorescence, the peak temperature of thermoluminescence bands, and rates of oxygen evolution, indicate little perturbation to electron transfer through the PSII complex. The susceptibility of mutant PSII to photoinactivation as measured by the light-induced loss of PSII activity in whole cells in the presence of the protein-synthesis inhibitors chloramphenicol or lincomycin was similar to that of wild type. These results indicate that phosphorylation at Thr-2 is not required for PSII function or for protection from photoinactivation. In control experiments the phosphorylation of D2 in wild-type C. reinhardtii was examined by 32P labeling in vivo and in vitro. No evidence for the phosphorylation of D2 in the wild type could be obtained. [14C]Acetate-labeling experiments in the presence of an inhibitor of cytoplasmic protein synthesis also failed to identify phosphorylated (D2.1) and nonphosphorylated (D2.2) forms of D2 upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Our results suggest that the existence of D2 phosphorylation in C. reinhardtii is still in question.

The 9-kDa phosphoprotein of photosystem II. Generation and characterisation of Chlamydomonas mutants lacking PSII-H and a site-directed mutant lacking the phosphorylation site.

The chloroplast gene psbH encodes a 9-10 kDa thylakoid membrane protein (PSII-H) that is associated with photosystem II and is subject to light-dependent phosphorylation at a threonine residue located on the stromal side of the membrane. The function of PSII-H is not known, neither is it clear what regulatory role phosphorylation may play in the control of PSII activity. Using particle gun-mediated transformation, we have created chloroplast transformants of Chlamydomonas reinhardtii in which the synthesis of PSII-H is prevented by the disruption of psbH, or in which the phosphorylatable threonine is replaced by alanine through site-directed mutagenesis of the gene. The mutants lacking PSII-H have a photosystem II-deficient phenotype, with no detectable functioning PSII complex present in whole cells or isolated thylakoid membranes. In contrast, the alanine mutant (T3A) grows photoautotrophically, and PSII activity is comparable to wild-type cells as determined by various biochemical and biophysical assays.

Reduced turnover of the D1 polypeptide and photoactivation of electron transfer in novel herbicide resistant mutants of Synechocystis sp. PCC 6803.

Two missense mutants, A263P and S264P, and a deletion mutant des-Ala263, Ser264, have been constructed in the D1 protein of the cyanobacterium Synechocystis sp PCC 6803. All were expected to induce a significant conformational change in the QB-binding region of photosystem II (PSII). Although the des-Ala263, Ser264-D1 mutant accumulated some D1 protein in the thylakoid membrane it was unable to grow photoautotrophically or evolve oxygen. Thermoluminescence and chlorophyll fluorescence studies confirmed that this deletion mutant did not show any functional PSII activity. In contrast, [S264P]D1 was able to grow photoautotrophically and give light-saturated rates of oxygen evolution at 60% of the rate of the wild-type control strain, TC31. The A263P missense mutant was also able to evolve oxygen at 50% of TC31 rates although it did not readily grow photoautotrophically. Thermoluminescence, flash oxygen yield and chlorophyll fluorescence measurements indicated that in both missense mutants electron transfer from QA to QB was significantly impaired in dark adapted cells. However, QA to QB electron transfer could be photoactivated in the mutants by background illumination. Both the A263P and S264P mutants also showed an increase in resistance to the s-triazine family of herbicides although this feature did not hold for the phenolic herbicide, ioxynil. Of particular interest was that the two missense mutants, especially S264P, possessed a slower rate of turnover of the D1 protein compared with TC31 and in vivo contained detectable levels of a 41-kDa adduct consisting of D1 and the alpha subunit of cytochrome b559. When protein synthesis was blocked by the addition of lincomycin, D1 degradation was again slower in S264P than TC31. The results are discussed in terms of structural changes in the QB-binding region which affect herbicide and plastoquinone binding and perturb the normal regulatory factors that control the degradation of the D1 protein and its synchronisation with the synthesis of a replacement D1 protein.

The lumenal loop connecting transmembrane helices I and II of the D1 polypeptide is important for assembly of the photosystem two complex.

Current structural models indicate that the D1 and D2 polypeptides of the Photosystem two reaction center complex (PS II RC) each span the thylakoid membrane five times. In order to assess the importance of the lumenal extrinsic loop that connects transmembrane helices I and II of D1 we have constructed five deletion mutants and two double mutants in the cyanobaterium Synechocystic sp. PCC 6803. Four of the deletion mutants (Δ59-65, Δ69-74, Δ79-86 and Δ109-110) are obligate photoheterotrophs unable to accumulate D1 in the membrane as assayed by immunoblotting experiments or pulse-labelling experiments using [(35)S]-methionine. In contrast deletion mutant Δ100 which lacks A100 behaved very similarly to the WT control strain in terms of photoautotrophic growth rate, saturated rates of oxygen evolution, flash-induced oxygen evolution, fluorescence induction and decay, and thermoluminescence. Δ100 is the first example of an internal deletion on the lumenal side of the D1 polypeptide that is benign to photosystem two function. Double mutant D103G/E104A also behaves similarly to the WT control strain leading to the conclusion that residues D103 and E104 are unlikely to be involved in ligating the metal ions Mn or Ca(2+), which are needed for photosynthetic oxygen evolution. Double mutant, G109A/G110A, was constructed to assess the significance of this GlyGly motif which is also conserved in the L subunit of purple bacterial reaction centres. The G109A/G110A mutant is able to evolve oxygen at approximately 50-70% of WT rates but is unable to grow phatoautotrophically apparently because of an enhanced sensitivity to photoinactivation than the WT control strain. A photoautotropic revertant was isolated from this strain and shown to result from a mutation that restored the WT codon at position 109. Pulse-chase experiments in cells using [(35)S]-methionine showed that resistance to photoinhibition in the revertant correlated with an enhanced rate of incorporation of D1 into the membrane compared to mutant G109A/G110A. The sensitivity to photoinhibition shown by the G109A/G110A mutant is therefore consistent with a perturbation to the D1 repair cycle possibly at the level of D1 synthesis or incorporation of D1 into the PS II complex.

Deletion of the PEST-like region of photosystem two modifies the QB-binding pocket but does not prevent rapid turnover of D1.

The rapid turn-over of the D1 polypeptide of the photosystem two complex has been suggested to be due to the presence of a "PEST"-like sequence located between putative transmembrane helices IV and V of D1 (Greenberg, B. M., Gaba, V., Mattoo, A. K. and Edelman, M. (1987) EMBO J. 6, 2865-2869). We have tested this hypothesis by constructing a deletion mutant (delta 226-233) of the cyanobacterium Synechocystis sp. PCC 6803 in which residues 226-233 of the D1 polypeptide, containing the PEST-like sequence, have been removed. The resulting mutant, delta PEST, is able to grow photoautotrophically and give light-saturated rates of oxygen at wild type levels. However electron transfer on the acceptor side of the complex is perturbed. Analysis of cells by thermoluminescence and by monitoring the decay in quantum yield of variable fluorescence following saturating flash excitation indicates that Q-B, but not Q-A, is destabilized in this mutant. Electron transfer on the donor side of photosystem two remains largely unchanged in the mutant. Turnover of the D1 polypeptide as examined by pulse-chase experiments using [35S]methionine was enhanced in the delta PEST mutant compared to strain TC31 which is the wild type control. We conclude that the PEST sequence is not absolutely required for turnover of the D1 polypeptide in vivo although deletion of residues 226-233 does have an effect on the redox equilibrium between QA and QB.

Flash oxygen yield patterns of autotrophically and photoheterotrophically grown Chlamydobotrys stellata in the presence and absence of lipophilic acceptors.

The obligate phototrophic green alga Chlamydobotrys stellata does not evolve oxygen when grown in CO2-free atmosphere on acetate. With the application of the lipophilic acceptor 2,6-dichloro-p-benzoquinone it was investigated whether this phenomenon is caused by the inactivation of the water-splitting system or by an inhibition of the electron transport chain. It was found that in the presence of DCQ, the photoheterotrophic alga exhibited a normal period-4 flash oxygen pattern, but the steady state yield was only 25% of that measured in the autotrophic cells. After DCQ addition, the initial distribution of S-states and the values of the transition probabilities proved to be the same in the autotrophic and photoheterotrophic algae. These results indicate that photoheterotrophic growth conditions inhibit the electron transport of Chl. stellata behind the acceptor site of DCQ, but the water-splitting system remains active with a reduced oxygen evolving capacity.

AS-1 cyanophage infection inhibits the photosynthetic electron flow of photosystem II in Synechococcus sp. PCC 6301, a cyanobacterium.

In Synechococcus sp. cells AS-1 cyanophage infection gradually inhibits the photosystem II mediated photosynthetic electron flow whereas the activity of photosystem I is apparently unaffected by the cyanophage infection. Transient fluorescence induction and flash-induced delayed luminescence decay studies revealed that the inhibition may occur at the level of the secondary acceptor, QB of photosystem II. In addition, the breakdown of D1-protein is inhibited, comparable to DCMU-induced protection of D1-protein turnover, in AS-1-infected cells.