Sequencing, cloning and expression of a beta-1,4-mannanase gene, manA, from the extremely thermophilic anaerobic bacterium, Caldicellulosiruptor Rt8B.4.

A gene encoding a beta-mannanase (manA) has been cloned from an obligately anaerobic extreme thermophile, Caldicellulosiruptor strain Rt8B.4, which is most closely related to Caldicellulosiruptor saccharolyticus (formerly Caldocellum saccharolyticum). The gene codes for a multidomain enzyme with a C-terminal beta-mannanase domain which was amplified by the polymerase chain reaction and cloned into a temperature-inducible expression vector in Escherichia coli. Sequence comparisons have shown that the Man domain of Rt8B.4 ManA is related to a thermophilic Dictyoglomus mannanase and a mesophilic mannanase from a Bacillus species. It appears to be unrelated to the beta-mannanase domain of C. saccharolyticus, implying acquisition of the genes from unrelated sources by the two bacteria.

Cervical ripening in humans: potential roles of estrogen, progesterone, and insulin-like growth factor-I.

OBJECTIVE: During pregnancy in humans a gradual connective tissue remodeling takes place in the cervix. The aim of this study was to examine a possible relationship between the action of gonadal steroids and growth factors and the biochemically identifiable changes in connective tissues during cervical ripening. STUDY DESIGN: Cervical biopsy specimens and serum samples were taken from 20 term pregnant and 20 nonpregnant menstruating women. Estrogen receptors and progesterone receptors were measured with enzyme immunoassays. The messenger ribonucleic acid levels for estrogen receptors, progesterone receptors, and insulin-like growth factor-I were determined by solution hybridization with human complementary deoxyribonucleic acid probes. The concentration of collagen and its solubility by pepsin digestion were measured. Statistical evaluations were done with the Student t test. RESULTS: In term pregnancy the estrogen receptor level decreased to 14% and the progesterone receptor level to 24% of nonpregnant levels (p <0.001 and p <0.01). The insulin-like growth factor-I messenger ribonucleic acid level increased 400% (p <0.01), whereas the messenger ribonucleic acid levels for estrogen receptors and progesterone receptors were unchanged. The changes coincided with a twofold decrease in collagen concentration (hydroxyproline) and a twofold increase in collagen solubility. CONCLUSION: Estrogen receptors and progesterone receptors are present in human cervix. A significant down-regulation of estrogen receptors and progesterone receptors and a fourfold increase in the insulin-like growth factor-I messenger ribonucleic acid level were registered in term pregnant cervix. These findings coincided with the remodeling of the cervical connective tissue.