Sulfmyoglobin. Resonance Raman spectroscopic evidence for an iron-chlorin prosthetic group

The green heme protein sulfmyoglobin (SMb) has been suggested to contain a sulfur-modified iron chlorin prosthetic group. To evaluate this hypothesis, we have obtained high-frequency (greater than 1000 cm-1) resonance Raman spectra of both oxidized and reduced SMb with 457.9-, 488.0-, 514.5-, 568.2-, and 647.1-nm excitation. The SMb spectra are compared to those of native met- and deoxymyoglobin (Mb). Vibrational frequencies for SMb are generally similar to those of Mb, suggesting a high-spin state for both the Fe(III) and Fe(II) SMb species, as is typical of native Mb. However, major differences between SMb and Mb occur both for patterns of relative spectral intensities and for depolarization ratios. In particular, all B1g-depolarized porphyrin modes in the Mb spectra have become polarized, totally symmetric vibrational modes in the SMb spectra. These contrasts reflect a dramatic lowering of the effective symmetry for the SMb prosthetic group. Several new bands are observed in SMb spectra that are not present in spectra of either native Mb or iron protoporphyrin IX complexes. The observation of additional polarized bands flanking the oxidation state marker, V4, is of particular interest. In a parallel study, we compared the resonance Raman spectral properties of iron protoporphyrin IX-derived chlorins and metallo-octaethylchlorins with those of the analogous porphyrins: the chlorin spectra exhibited altered intensity patterns, an increased number of totally symmetric (polarized) vibrational bands, and several new vibrational bands, including one or two in the region of the oxidation state marker, V4. Thus, the resonance Raman spectral characteristics of SMb and metallo-chlorins are complementary and strongly support a chlorin prosthetic group for SMb. Furthermore, they establish testable criteria for investigating the prosthetic group structures of other green heme proteins by resonance Raman spectroscopy.     

大鼠非致瘤四倍体细胞系的建立及细胞遗传学和酶学特征

从大鼠自然诱发的肉瘤细胞中,我们建立了一个四倍体细胞系(4n=84),命名为RC(ratcell)。它具有典型的成纤维细胞外形,能在玻璃表面贴壁生长,但不能生长在琼脂半固体培养基中。该细胞在含15％小牛血清的RPMI 1640培养基中生长良好,至今已连续繁殖112世代,细胞群体倍增时间约为15小时。染色体G-带分析表明,RC为整四倍体细胞,它的1条X染色体在第32至34区为均染区。RC细胞核仁组织者(NORs)活性显然比大鼠二倍体细胞NORs活性的加倍还高(P<0.001)。这个具有非常高NORs活性的RC细胞系对于研究细胞18S+28S rRNA基因转录活性的调控、基因表达与基因剂量关系有一定的意义。RC细胞还有异常高的磷酸酯酶活性,而且它的同工酶谱也与大鼠肌肉细胞明显不同。体内接种实验和扫描电镜的观察表明,RC是非致瘤细胞。RC细胞各号染色体的C-带图样与大鼠二倍体细胞无明显的差异。     

Influence of both coculture and BrdU on NOR activity of mouse,rat and human cells

Summary The coculture of mouse PG19 cells with human MGC cells can significantly suppress nucleolar organizer region (NORs) activity of both PG19 and MGC cells. 5-bormodeoxyuridine (BrdU) can also significantly suppress the NOR activity of rat RC cells, human MGC and Hela cells, and mouse PG19 cells: i.e. the average number of Ag-NORs and the number of chromosomes bearing Ag-NORs per cell decrease significantly. The degree of the suppression increases with increase in both BrdU concentration in the culture medium and BrdU treatment time. The suppressed NOR activity of the PG19 cells can gradually be restored when the BrdU-treated cells are transferred into BrdU-free medium for 50 h. In PG19 cells deoxycytidine (dC) can reverse the suppression of NOR activity caused by BrdU. Coculture plus BrdU treatment suppress the NOR activity of PG19 cells more severely than BrdU treatment alone. In coculture medium containing 30 g BrdU/ml, dC can also reverse the suppression of the NOR activity of PG19 cells but not that of the MGC cells. The degree of the reversion in the coculture plus BrdU treatment is significantly lower than that found with BrdU-treatment alone.     

High-performance liquid chromatography of uroporphyrinogen and coproporphyrinogen isomers with amperometric detection.

A reversed-phase h.p.l.c. system, with an ODS-Hypersil column with acetonitrile or methanol in ammonium acetate buffer as mobile phase, is described for the separation of uro-and copro-porphyrinogen isomers. The porphyrinogens are detected amperometrically with sensitivity comparable with that of the fluorescent detection of porphyrins. The effects of pH, buffer concentration and organic modifiers on retention and resolution were studied. The method is suitable for both analytical and preparative separation of porphyrinogens.     

The effect of 1-β-d-arabinofuranosyl-cytosine on the expression of the common fragile site at 3p14

Summary The effect of the G₂-treatment of 1--d-arabino-furanosyl-cytosine (araC) on the expression of the common fragile site at 3p14 (FRA3B) was studied. A significantly increased frequency of FRA3B induced by G₂ treatment of araC was found in the lymphocytes grown in folate-deficient medium (positive rate 100%). A relatively low frequency of FRA3B was also induced in the cultures with folate in four of the seven subjects. These is a synergistic effect between araC and growth in folate-deficient medium on the induction of FRA3B. The results suggest that the DNA lesions related to the expression of FRA3B induce the long-patch repair and that the low DNA polymerase activity and inefficient repair process during G₂ phase is involved in the expression of FRA3B.