Rubisco condensate formation by CcmM in ß-carboxysome biogenesis.

Cells use compartmentalization of enzymes as a strategy to regulate metabolic pathways and increase their efficiency1. The α- and ß-carboxysomes of cyanobacteria contain ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco)-a complex of eight large (RbcL) and eight small (RbcS) subunits-and carbonic anhydrase2-4. As HCO3- can diffuse through the proteinaceous carboxysome shell but CO2 cannot5, carbonic anhydrase generates high concentrations of CO2 for carbon fixation by Rubisco6. The shell also prevents access to reducing agents, generating an oxidizing environment7-9. The formation of ß-carboxysomes involves the aggregation of Rubisco by the protein CcmM10, which exists in two forms: full-length CcmM (M58 in Synechococcus elongatus PCC7942), which contains a carbonic anhydrase-like domain8 followed by three Rubisco small subunit-like (SSUL) modules connected by flexible linkers; and M35, which lacks the carbonic anhydrase-like domain11. It has long been speculated that the SSUL modules interact with Rubisco by replacing RbcS2-4. Here we have reconstituted the Rubisco-CcmM complex and solved its structure. Contrary to expectation, the SSUL modules do not replace RbcS, but bind close to the equatorial region of Rubisco between RbcL dimers, linking Rubisco molecules and inducing phase separation into a liquid-like matrix. Disulfide bond formation in SSUL increases the network flexibility and is required for carboxysome function in vivo. Notably, the formation of the liquid-like condensate of Rubisco is mediated by dynamic interactions with the SSUL domains, rather than by low-complexity sequences, which typically mediate liquid-liquid phase separation in eukaryotes12,13. Indeed, within the pyrenoids of eukaryotic algae, the functional homologues of carboxysomes, Rubisco adopts a liquid-like state by interacting with the intrinsically disordered protein EPYC114. Understanding carboxysome biogenesis will be important for efforts to engineer CO2-concentrating mechanisms in plants15-19.

Plant RuBisCo assembly in E. coli with five chloroplast chaperones including BSD2.

Plant RuBisCo, a complex of eight large and eight small subunits, catalyzes the fixation of CO2 in photosynthesis. The low catalytic efficiency of RuBisCo provides strong motivation to reengineer the enzyme with the goal of increasing crop yields. However, genetic manipulation has been hampered by the failure to express plant RuBisCo in a bacterial host. We achieved the functional expression of Arabidopsis thaliana RuBisCo in Escherichia coli by coexpressing multiple chloroplast chaperones. These include the chaperonins Cpn60/Cpn20, RuBisCo accumulation factors 1 and 2, RbcX, and bundle-sheath defective-2 (BSD2). Our structural and functional analysis revealed the role of BSD2 in stabilizing an end-state assembly intermediate of eight RuBisCo large subunits until the small subunits become available. The ability to produce plant RuBisCo recombinantly will facilitate efforts to improve the enzyme through mutagenesis.

Iodine supplementation in Austria: methods and results.

Until 1963 Austria was an extremely iodine-deficient area with low iodine intake and high goiter prevalence. Therefore, for the first time in 1963, salt iodination with 10 mg of potassium iodide per kilogram of salt was introduced by federal law. Twenty years after this salt iodination, however, investigations in schoolchildren demonstrated iodine deficiency grade I to II according to the World Health Organization (WHO) (urinary iodine excretion, 42-75 microg/g Crea) and goiter prevalence of far more than 10%. In 1990, salt iodination was increased to 20 mg of potassium iodide per kilogram of salt. In 1994, further investigations in schoolchildren demonstrated an increase of urinary iodine excretion (121 microg/g Crea) and a reduction of goiter prevalence below 5%, with the exception of pupils ages 14-19 (12%). In the year 2000, 10 years after the increase of salt iodination in Austria, 430 nonselected adult inhabitants of three communities in Carinthia (a county of Austria) were investigated for iodine excretion, goiter prevalence, and prevalence of thyroid autoantibodies. This study demonstrated that although iodine supply is sufficient now in Austria (males, 163.7 microg of Crea; females, 183.3 microg of iodine per gram of Crea), goiter prevalence is still high in the elderly, who lived for a longer period of iodine deficiency (34.3% in women and 21.3% in men), whereas goiter prevalence in younger people up to age 40 years is below 5%. It could also be shown that the percentage of thyroid autoantibodies is now as high as in other countries with sufficient iodine supply (3.19% in males, 5.17% in females). In addition to the changes of urinary iodine excretion and goiter prevalence because of salt iodination, changes of incidence in hyperthyroidism and histologic types of thyroid cancer are discussed in this paper. In conclusion, the introduction of salt iodination led to an improvement in iodine supply with a marked reduction of goiter prevalence in people who were born after 1963, but also to an increase in hyperthyroidism and autoimmune thyroid diseases as well as changes in histologic types of thyroid cancer.

Study of the effect of gamma-irradiation on bovine serum samples on the ability of monoclonal antibodies to detect invariant antigens of Trypanosoma congolense, T. vivax and T. brucei in enzyme-linked immunosorbent assays.

Samples of bovine serum from uninfected and African trypanosomes-infected animals were tested before and after gamma-irradiation, using three sandwich enzyme-linked immunosorbent assays (ELISA). Each test system utilized a different monoclonal antibody, reputedly allowing the specific detection of conserved-invariant cytoplasmic antigens of trypanonosomes, T. congolense, T. vivax, and T. brucei, respectively. Results have identified two groups of samples. The first contained samples where there were unequivocal ELISA results indicating positivity and negativity, for non-irradiated samples. In this group, irradiation had no effect on the diagnostic sensitivity of the assays. All samples shown to be positive before irradiation remained positive and those shown to be negative, remained negative. There was, however, a statistically significant reduction in signal in each of the ELISAs following irradiation. The second group contained samples identified before irradiation as flanking the diagnostic negative/positive threshold of OD > or =0.05. These showed a negative bias after irradiation of the order of OD -0.01, which was shown to be statistically significant by paired t-statistics. Without correction of the given diagnostic negative/positive threshold, bovine sera with OD values around the threshold were expected to deliver more false negative test results upon irradiation. This was confirmed when serological data were compared with parasitological findings; where three times more false negative test results were found from irradiated serum samples. Consequently, for this group of irradiated bovine samples tested by ELISA, the re-adjustment of the diagnostic negative/positive threshold of the ELISAs using defined irradiated serum samples is recommended; otherwise, the frequency of false negative results might be increased.

[Long-term laryngoscopic follow-up in vocal cord paralysis following struma surgery]. / Die langfristige laryngoskopische Nachkontrolle bei Einschränkung der Stimmbandmotilität nach Strumaoperation.

Of 525 patients 17 (3.2%) showed a laryngoscopically established palsy of the recurrent laryngeal nerve after surgery due to goiter. A laryngoscopic follow-up of all these patients, performed at least one year after the operation, revealed that 76.5% of the recurrent nerve palsies were temporary and 23.5% were permanent. Danger of permanent palsy increased in the sequence--uncomplicated nodular goiter--struma maligna--recurrent goiter. The outcome of long-term follow-up showed a palsy rate of 0.8%, which was much lower than the corresponding rate reported by short-term control (p = 0.005). Therefore laryngoscopic long-term follow-up in cases of postoperative abnormal laryngoscopic function should be a standard part of follow-up in thyroid gland surgery.

[Carotid artery surgery with local anesthesia. Experiences with a series of 181 operations]. / Karotischirurgie in Lokalanästhesie. Erfahrungen mit einer Serie von 181 Operationen.

Between 1982 and 1984, 181 operations of the carotid artery were performed--180 of these in primary awake patients--36 in stage I (19.9%), 72 in stage II (39.8%), 72 in stage IV (39.8%) and only one in stage III (0.6%). 144 patients (79.6%) tolerated clamping of the carotid artery very well. In 37 patients (20.4%) clamping was not tolerated (two patients, in whom the control of cerebral function was not possible, were submitted to the latter group). In three of 181 patients clamping-intolerance started at the end of operation, more than 30 minutes after clamping. Clamping intolerance in these cases was found to be confined to pulmonary and cardiac factors, no shunt was used, but operation was brought to an end as quickly as possible. Complications were present in 15 patients (8.3%), three of them resulting in death (hospital mortality rate 1.7%), 2 of them in permanent loss of function (morbidity rate 1.1%). Only in 2 patients (asystolism with consecutive successful reanimation in one case, and thoracotomy due to enlargement of surgical procedure in the other case) a primary general anaesthesia would have facilitated surgery. Surgery of the carotid artery under local anaesthesia is considered to be a guarantee for a secure intraoperative surveillance followed by a low postoperative complication rate. In a few cases more difficult surgical procedures have to be accepted in account of that.

[Temporary and permanent recurrent laryngeal nerve paralysis following thyroid surgery. Modifying factors: the educational status of the surgeon]. / Die passagere und permanente Rekurrenzparese nach Schilddrüsen-operationen. Beeinflussende Faktoren: der Ausbildungsstand des Chirurgen.

17 of 525 patients (3.2%) showed an laryngoscopically established palsy of the recurrent laryngeal nerve after surgery for struma. The analysis of these operations, performed by five surgeons during or within three years after the period of surgical training, revealed that the operations performed under assistance of the senior surgeons were high grade selected (p = 0.026). Thus 14.8% of the operations performed because of simple goiter but only 4.8% of the operations performed because of thyroid cancer/recurrent goiter/extensive nodular goiter were assisted in this way. On the other hand it was necessary to call for help of a senior surgeon because of intraoperative difficulties in only 1.26% of the cases operated on for simple goiter, but in 19.6% of the more complex forms of goiter (p less than 0.001). The risk of recurrent laryngeal nerve palsy was nearly 10 times higher in the complex forms of goiter than in the simple forms (p less than 0.001). More extensive surgical training in the forms of complex goiters should be able to improve the results.

[Comparison between the biochemical and histochemical determination of hormone receptors in breast carcinomas]. / Vergleich zwischen biochemischer und histochemischer Hormonrezeptorenbestimmung bei Mammakarzinomen.

The hormonal sensitivity of breast cancer on one hand indicates the prognosis and on the other determines the therapeutical procedure when metastases appear. It also seems to be important in the choice of adjuvant therapy. Therefore the analysis of hormonal receptors in our opinion is at least as important as the histological typification of the carcinoma. As there was a trend recently showing up the so-called histochemical steroid-receptor-assay which is much less complicated than the biochemical (quantitative) method, we investigated the value of this fluorescence-microscopical assay by means of parallel determinations. In the results of 157 patients of the last three years we found differences in 45% of all oestrogen-receptor-determinations and in 50% of 29 progesterone-receptor-determinations. This indicates that the histochemical assay is most probably not representative. There is serious doubt about the possibility to document the hormone receptor by this method at all. Therefore it does not seem advisable at the moment to waive the biochemical receptor determination.