Import of a new chloroplast inner envelope protein is greatly stimulated by potassium phosphate

A cDNA clone encoding a major chloroplast inner envelope membrane protein of 96 kDa (IEP96) was isolated and characterized. The protein is synthesized as a larger-molecular-weight precursor (pIEP96) which contains a cleavable N-terminal transit sequence of 50 amino acids. The transit peptide exhibits typical stromal targeting information. It is cleaved in vitro by the stromal processing peptidase, though the mature protein is clearly localized in the inner envelope membrane. Translocation of pIEP96 into chloroplasts is greatly stimulated in the presence of 80 mM potassium phosphate which results in an import efficiency of about 90%. This effect is specific for potassium and phosphate, but cannot be ascribed to a membrane potential across the inner envelope membrane. Protein sequence analysis reveals five stretches of repeats of 26 amino acids in length. The N-terminal 300 amino acids are 45% identical (76% similarity) to the 35 kDa -subunit of acetyl-CoA carboxyl-transferase from Escherichia coli. The C-terminal 500 amino acids share significant similarity (69%) with USOI, a component of the cytoskeleton in yeast.Abbreviations P_i phosphate - IEP inner envelope membrane protein - pIEP precursor form of IEP - SSU small subunit of ribulose-1,5-bisphosphate carboxylase oxygenase - IEP96_pep peptide specific antiserum to IEP96 - IEP96_pol polyspecific antiserum to IEP96     

Fossil Homo femur from Berg Aukas,northern Namibia

The proximal half of a hominid femur was recovered from deep within a paleokarst feature at the Berg Aukas mine, northern Namibia. The femur is fully mineralized, but it is not possible to place it in geochrono logical context. It has a very large head, an exceptionally thick diaphyseal cortex, and a very low collodiaphyseal angle, which serve to differentiate it from Holocene homologues. The femur is not attributable to Australopithecus, Paranthropus, or early Homo (i.e., H. habilis sensu lato). Homo erectus femora have a relatively longer and AP flatter neck, and a shaft that exhibits less pilaster than the Berg Aukas specimen. Berg Aukas also differs from early modern femora in several features, including diaphyseal cortical thickness and the degree of subtrochanteric AP flattening. The massive diaphyseal cortex of Berg Aukas finds its closest similarity within archaic H. sapiens (e.g., Castel di Guido) and H. erectus (e.g., KNM-ER 736) samples. It has more cortical bone at midshaft than any other specimen, although relative cortical thickness and the asymmetry of its cross-sectional disposition at this level are comparable with those of other Pleistocene fem ora. The closest morphological comparisons with Berg Aukas are in archaic (i.e., Middle Pleistocene) H. sapiens and Neandertal samples. © 1995 Wiley-Liss, Inc.     

Structural and conformational modifications of alpha-MSH/ACTH4-10 provide melanotropin analogues with highly potent behavioral activities

Previous studies have identified the (4-10) heptapeptide sequence as the central core of alpha-MSH/ACTH peptides required for mediation of important biological activities. In the present study, the structure-activity relationships of Nle4-substituted and Cys4,Cys10-bridged cyclic alpha-MSH analogues, which were previously shown to exhibit a wide range of melanotropic potencies from weak agonism to super potency, were examined for grooming behavioral activity in the rat following intracerebroventricular injections. The results showed that stepwise C-terminal elongation of the linear Nle4-substituted Ac-alpha-MSH4-10-NH2 increased grooming potencies of the peptides in a manner similar to their actions on melanocytes. The most interesting finding was the observation that cyclization of the inactive linear "central (4-10) core" of alpha-MSH (Ac-alpha-MSH4-10) to form Ac-[Cys4,Cys10]-alpha-MSH4-10-NH2 resulted in a super potent agonist in the grooming assay. However, while cyclization of the (4-10) heptapeptide produced potent agonists on grooming behavior, the structure-activity relationships were different than the frog skin bioassay. These findings support the hypothesis that appropriate structural and confirmational modifications of alpha-MSH-related peptides can produce profound effects on the bioactivities of the peptides, and suggest that different structural-conformational requirements exist for alpha-MSH interactions with its various receptors.     

Regulation of the host response to bacterial lipopolysaccharides

Bacterial endotoxins or lipopolysaccharides (LPS) are unique glycolipids present in the outer cell membrane of all gram-negative bacteria. It is now generally recognized that LPS is of primary importance in initiating the pathophysiological changes that often accompany gram-negative bacillary infections in humans including hypotensive shock, disseminated intravascular coagulation, and metabolic abnormalities. Although the biochemical mechanisms of these changes are not well understood, increasing emphasis has been placed on defining the biochemical response of the macrophage (M phi) to LPS. In this paper we describe two M phi-derived factors induced by LPS that may be important in the expression of endotoxic activity in the host. These are a procoagulant activity, which is present on the cell membrane of LPS-treated rabbit liver M phi and acts by directly activating coagulation factor X, and a factor released into the supernatant by LPS-treated peritoneal exudate M phi, which suppresses steroidogenesis in explanted adrenocortical cells. The potential role of the M phi in regulating the binding of LPS to high-density lipoproteins through the induction of acute phase proteins is also considered.     

Estradiol 17β-dehydrogenase and 20α-hydroxysteroid dehydrogenase from human placental cytosol: one enzyme with two activities?

The soluble enzyme estradiol 17β-dehydrogenase (17β-ED) from human term placental cytosol is reported to be a stereospecific oxidoreductase for estrogen substrates. A published purification scheme (heat treatment and affinity chromatography) yielded a homogeneous protein which had the reported characteristics of pure 17β-ED and also had 20α-hydroxysteroid dehydrogenase (20α-HSD) activity. Spectrophotometric assay when the buffer contained albumin, 8 mg/ml, masked the 20α-HSD activity observed in albumin-free conditions and may explain why this bifunctional activity has gone unrecognized. In human placenta, one enzyme may catalyze stereospecific oxidation/reduction of both estrogen and progesterone.     

Restriction analysis of the Streptomyces glaucescens genome by agarose gel electrophoresis

A method for the analysis of total DNA of Streptomyces glaucescens is described. The relevant steps are (a) extraction and purification of DNA, (b) restriction of DNA samples with type II restriction enzymes, (c) one dimensional separation of restriction fragments by agarose gel electrophoresis. A typical banding pattern was obtained for each wild type strain, independant of growth conditions or age of the culture. Mutant strains exhibited in most cases the same banding pattern as the parent wild type strain. Only in one specific mutant class a fragment of about 9 megadalton was missing.     

The natural history of the helicoidal occlusal plane and its evolution in early Homo

In modern man the pitch of the occlusal plane may vary along the tooth-row. When anterior cheek-teeth show a plane sloping upward palatally, whilst that on posterior cheek-teeth slopes upward buccally, there results a twisted or helicoidal occlusal plane (Ackermann). Several hypotheses have been proposed for the structural basis of the helicoidal occlusal plane. Campbell's proposal ('25) has gained widest acceptance, namely that the helicoid results from anteroposterior differences in upper and lower alveolar arch width. In the early 1960s, while studying the Olduvai hominids assigned to Homo habilis, the author noted changing occlusal slopes along the tooth-row and a slight helicoid, although these featues had not been noted in other early hominids. Subsequently, Wallace showed a total absence of the helicoid from South African australopithecines, and its presence in Swartkrans Homo, SK 45 and SK 80. Recent studies confirm the presence of the helicoid in all available specimens of H. habilis, including Stw 53 found at Sterkfontein in 1976. Hence, this trait may distinguish between Australopithecus and early Homo. Measurements of the maxillary arch widths have shown that, whereas in Australopithecus arch widths increase to a maximum at M3, in early Homo maxillary arch widths are greatest at M2. The decline in posterior maxillary arch width is part of a general reduction of that region. Thus despite striking elongation of premolars and M1 in early Homo, M2 and M3 are mesiodistally abbreviated. It is hypothesized that the onset of posterior arch reduction, with the appearance of a helicoid, was a structural and functional concomitant of the transition from the presumed australopithecine ancestor to H. habilis.     

Trace metal modification of immunocompetence. I. Effect of trace metals in the cultures on in vitro transformation of B lymphocytes.

Lymphocytes obtained from the spleen of Balb/C mice have been subjected to transformation by LPS in the presence of varying concentrations of Pb²⁺, Cd²⁺, or Cr³⁺. Both DNA and protein turnover were followed. It was found that Pb²⁺ and Cr³⁺ are mitogenic over a broad range of concentrations, while Cd²⁺ is slightly mitogenic at very low (10^?6M) concentration and rapidly becomes inhibitory of both [³H]TdR and [³H]ALA uptake. Pb²⁺ appears to stimulate the action of LPS, while Cr³⁺ appears to inhibit. Each of the metals protects the lymphocytes from cell death arising from incubation with LPS. The mechanism of the observed changes is as yet obscure.