Objective To express insulin-degrading enzyme(IDE)mutant T142A in prokaryotic cells and detect its activity.Methods According to the results of multi-sequence alignment and IDE substrate co-crystal structure,an active residue in β6-strand structure of IDE were predicted.The recombinant plasmid ppSUMO-T142A,with the site mutation of threonine 142 to alanine,was constructed by point mutation technique and expressed by E.coli prokaryotic expression system.After purification by nickel ion column affinity chromatography,ion exchange chromatography and gel filtration chromatography,the mutant T142A was obtained and determined for the activity by fluorescence method.Results IDE amino acid sequence is highly conserved among 16 species.T142 directly participates in substrate binding,interacts with substrate cleavage sites,and is close to important structures such as catalytic active sites and door-subdomains.The mutation of recombinant plasmid ppSUMO-T142A was proved to be correct by sequencing.The expressed fusion protein His-SUMO-T142A was mainly existed in soluble form in the supernatant at a concentration of 18 mg/mL,with a relative molecular mass of about 131 000;After three steps of purification,the purity of mutant T142A reached 86%.The maximum reaction rate(V_(max))of T142A catalytic degradation of fluorescent substrate V was 501.06 min~(-1) and the Michaelis constant(K_m) was 9.01μmol/L.Compared with wild-type IDE(V_(max) was 2 814.32 min~(-1),K_m was 11.93μmol/L),the activity of T142A decreased significantly.Conclusion The activity of IDE mutant T142A expressed in this study greatly decreases,while T142 is an important residue for IDE to play its enzymatic function,which provides an experimental basis for the development of new IDE activity regulatory molecules.

Cardiovascular Diseases ; Follistatin-Related Proteins ; physiology ; Humans

Objective: To study the chemical constituents of Artemisia anomala. Methods: Various chromatographic techniques were adopted to separate the constituents, and the spectroscopic analysis was used to identify their structures. Results: Seven compounds were isolated and identified as (4aS, 7S, 7aR)-methyl 7-hydroxy-7-methyl-1, 4a, 5, 6, 7, 7a-hexahydrocyclopenta [c] pyran-4- carboxylate (1), rehmaglutin D (2), (E)-6-hydroxy-2, 6-dimethylocta-2, 7-dienoic acid (3), chrysoeriol (4), luteolin (5), apigenin (6), and p-coumaric acid (7). Conclusion: Compound 1 is a new compound named artanoiridoid; Compound 2 is firstly reported in this genus; Compounds 3, 4, and 7 are separated from this plant for the first time.

The middle cerebral artery occlusion(MCAO) in rats remained a focus cerebral ischemic model that is non-incisioned,reliable,as well as the ischemic and refusion time can be controlled,which is widely used since it was created.However,because it is needed complex operations,associatied with many factors,it is difficult to establish.Many scholars have modified the operation,shape of the occlusion line,as well as the variety,narcotic drugs and so on.

Objective To explore the application of self-expanding stent and seperable detachable coil for interventional therapy in patients with wide-necked intracranial aneurysm.Methods 32 patients with wide-necked intracranial aneurysm received the interventional therapy by self-expanding stent and seperable detachable coil.After operation,the situation of the aneurysm occluded were observed by DSA immediately and followed up.Results The stents were successfully deployed in the 32 patients.The seperable detachable coils were occluded in the aneurysms in 29 cases.After operation immediately,the DSA showed that the aneurysms in 18 cases were completely occluded by seperable detachable coils,7 cases were 95% occluded,4 cases were 90% occluded.In the other 3 cases,the aneurysms were too small to be filled by seperable detachable coil,the stents were used only.All the parent arteries were no stenosis.18 h after operation,1 case died from multiple hemorrhage caused by thrombasthenia.Follow-up from 3 months to 1 year,DSA re-examination in 21 cases.Compared with the image immediately after operation,the imagings of DSA were no change in 12 cases,residual necks were disappeared in 5 cases and increased in 1 case.In the 3 cases which stents used only,the aneurysms were no development completely in the 2 cases and decreased in 1 case.6～18 months followed up in 26 cases,the rebleeding and cerebral infarction were no occurred.Conclusions The self-expanding stent is very flexible,secure,and effective.It can good support for the wide-necked aneurysms to be occluded by seperable detachable coils and effective promoting the small aneurysms healing.

Acupuncture Points ; Acupuncture Therapy ; instrumentation ; Aged ; Catgut ; Female ; Humans ; Male ; Middle Aged ; Muscle, Skeletal ; physiopathology ; Spasm ; physiopathology ; therapy ; Treatment Outcome

Humans ; Larynx ; surgery ; Neurilemmoma ; surgery

Blood Cells ; cytology ; Bone Marrow Cells ; cytology ; China ; Hematologic Tests ; standards ; Humans

Acute Disease ; Adolescent ; Adult ; Child ; Humans ; Leukemia ; diagnosis ; drug therapy ; Neoplasm, Residual ; Remission Induction

OBJECTIVE： To discuss the application prospect of pharmacogenomics METHODS： To introduce the concept of pharmacogenomics and analyse its application at prospect RESULTS ＆ CONCLUSION： Pharmacogenomics is wide in the prospect of application and has a vast reservoir of economic potential， it will exert an influence on pharmacy in the 21st century