The Rickettsia prowazekii citrate synthase (gitA) gene, previously cloned in Escherichia coli, was localized to a 2.0-kilobase chromosomnal fragment. DNA sequence analysis of a portion of this fragment revealed an open reading frame of 1,308 base pafrs that encodes a protein of 435 amino acids with a molecular weight of 49,171. This translation product is comparable in size to both the E. coli Snd pig heart citrate synthase monomers and to the protein synthesized in E. coli minicells containing the rickettsial gene. Comparisons between the deduced amino acid sequence of R. prowazekii citrate synthase and those of the E. coli and pig heart enzymes revealed extensive homology (59%) between the two bacterial proteins. In contrast, only 20% of the rickettsial enzyme residues were shared with the functionally similar pig heart enzyine residUes. Upstream from the open reading frame and in close proximity to one anothet, sequences with homology to E. coli consensus sequences for RNA polymerase and ribosome binding were identified. S1 nuclease mapping experiments demobstrated that the start of transcription for this gene in E. coli was located in the upstream region. Codon usage in the rickettsial git4 gene was found to be very biased and differed from the pattern observed in E. coli. Adenine and uracil were used preferentially in the third base position of rickettsial codons.Rickettsia prowazekii is an obligate intracellular parasitic bacterium that multiplies within the cytoplasm of its eucaryotic host cell rather than within a phagosome or phagolysosome. Its exploitation of this environment is reflected in a variety of novel features, most notably the existence of an ADP-ATP transport system that permits the rickettsiae to accumulate ATP directly from the cytoplasm of the host cell (44). However, the rickettsiae are not strict energy parasites and can generate their own ATP via the tricarboxylic acid cycle and oxidative phosphorylation (39). This ability is probably necessary for cell viability during periods when the host cell ATP reserves are depleted or during transit of the bacterium to a new host cell. Central to the regulation of the tricarboxylic acid cycle is the enzyme citrate synthase.Citrate synthase, the first enzyme of the tricarboxylic acid cycle, has been extensively studied in a variety of procaryotic and. eucaryotic cells (30,, 41, 43). These studies have identified two main types of citrate synthase: a "small" type (molecular weight, -106,000) found in eucaryotic cells and gram-positive bacteria and a "large" type (molecular weight, -250,000) found exclusively in gram-negative bacteria. The small enzyme is a dimer composed of two identical subunits, while the large enzyme is a multimer composed of four to six identical subunits. The two types of enzyme also differ in their sensitivity to certain effectors. The small enzyme is senlsitive in vitro to inhibition by ATP but not by at-ketoglutarate or NADH, while the large enzyme is sensitive in vitro to inhibition by a-ketoglutarate and NADH but not by ...

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This study tested the efficacy of assisted reproduction (synchronization of oestrus and intrauterine artificial insemination (AI)) in contributing to the captive propagation of an endangered species, the Eld's deer (Cervus eldi thamin). Semen was collected from males preselected on the basis of under-represented genotype. Motility of spermatozoa after thawing from ejaculates diluted with BF5F extender (8% glycerol), frozen on dry ice in 0.5 ml straws and stored in liquid nitrogen was 60-70%. Intravaginal progesterone-releasing devices (controlled internal drug release, CIDR-type G) were inserted into 20 adult Eld's deer hinds for 14 days. In all hinds, semen (7.5-10 x 10(6) motile spermatozoa per uterine horn) was deposited by laparoscopy performed 70 h after removal of the CIDR device. Ovarian activity, before and after AI, was monitored by analysing pregnanediol-3 alpha-glucuronide (PdG) concentrations in voided urine collected three to seven times per week. During the period of CIDR device insertion, urinary PdG profiles were equal to, or above, normal luteal phase concentrations in all hinds. Within 48 h of device withdrawal, PdG concentrations returned to baseline values in 17 of the 20 females, and the onset of behavioural oestrus occurred at this time in 12 hinds. On the basis of sustained increases in urinary PdG, 9 of the 20 hinds were diagnosed as pregnant by 90 days after AI, all of which delivered offspring after a mean gestation of 241.1 days (range, 235-245). Seven singletons (two females, five males) were born alive and survived, and one singleton and one set of twins were stillborn (three females).(ABSTRACT TRUNCATED AT 250 WORDS)

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