Packaging and unpackaging the sea urchin sperm genome.

Two species of histones in sea urchin sperm (Sp H1 and Sp H2B) are chimeric molecules whose highly basic amino-terminal domains are dephosphorylated at the last stage of sperm cell differentiation, and rephosphorylated immediately following fertilization. The phosphorylated regions consist largely of repeating tetrapeptides with two basic residues flanking Ser-Pro residues ('SPKK' motifs) and are predicted to have beta-turn secondary structures. Alteration of the charge and structure of the SPKK sites may play a role in the unusually dense DNA packaging of the mature sperm chromatin. The motif resembles the target site of cell-cycle-associated cdc2 kinases and is found in several other proteins whose nucleic acid affinities may be altered during the cell cycle.     

Asymmetric reconstitution of homogeneous Escherichia coli sn-glycerol-3-phosphate acyltransferase into phospholipid vesicles

The sn-glycerol-3-phosphate (glycerol-P) acyltransferase of Escherichia coli cytoplasmic membrane was purified in Triton X-100 (Green, P. R., Merrill, A. H., Jr., and Bell, R. M. (1981) J. Biol. Chem. 256, 11151-11159) and incorporated into mixed micelles containing Triton X-100, phosphatidylethanolamine, phosphatidylglycerol, cardiolipin, and beta-octyl glucoside. Enzyme activity was quantitatively reconstituted from the mixed micelle into single-walled phospholipid vesicles by chromatography over Sephadex G-50. Activity coeluted with vesicles of 90-nm average diameter on columns of Sepharose CL-4B and Sephacryl S-1000. These vesicles contained less than 2 Triton X-100 and 5 beta-octyl glucoside molecules/100 phospholipid molecules. Calculations suggested that up to eight 91,260-dalton glycerol-P acyltransferase polypeptides were incorporated per 90-nm vesicle. The pH dependence and apparent Km values for glycerol-P and palmitoyl-CoA of the glycerol-P acyltransferase reconstituted into vesicles were similar to those observed upon reconstitution by mixing of the enzyme in Triton X-100 with a 20-fold molar excess of sonicated phosphatidylethanolamine:phosphatidylglycerol:cardiolipin, 6:1:1. The integrity of vesicles containing glycerol-P acyltransferase was established by trapping 5,5'-dithiobis-(2-nitrobenzoic acid). Chymotrypsin inactivated greater than 95% of the glycerol-P acyltransferase in intact vesicles and cleaved the 91,260-dalton polypeptide into several vesicle-bound and several released peptides, indicating that critical domains of the enzyme are accessible in intact vesicles. Trinitrobenzene sulfonate and 4,4'-diisothiocyano-2,2'-disulfonic acid stilbene caused greater than 90% loss of glycerol-P acyltransferase in vesicles. Disruption of vesicles with Triton X-100 did not reveal significant latent activity. These data strongly suggest that the glycerol-P acyltransferase was reconstituted asymmetrically into the vesicles with its active site facing outward.     

Connective tissue activation. XXX: Isoelectric point microheterogeneity of CTAP-III, a human platelet-derived growth factor

Connective tissue activating peptide (CTAP-III) is one of the growth factors found in the alpha granules of human platelets. This small cationic platelet protein shows several apparent isoelectric point variants after the preparative isoelectric focusing step in large scale preparations from multiple donors. Analytical isoelectricfocusing techniques under denaturing conditions, coupled with Western blotting and immunodetection, now show that CTAP-III prepared rapidly from single donors also has multiple isoelectric point variants, suggesting that this finding is more likely related to post-translational modification than preparative artifact or genetic polymorphism.     

Effects of oral propranolol and exercise protocol on indices of aerobic function in normal man

The exercise responses to two different progressive, upright cycle ergometer tests were studied in nine healthy, young subjects either with no drug (ND) or following 48 h or oral propranolol (P) (40 mg q.i.d.). The ergometer tests increased work rate by 30 W either every 30 s or every 4 min. Propranolol caused a significant (p less than 0.05) reduction in peak oxygen uptake (VO2) during both the 30-s and 4-min tests (30-s ND, 3949 +/- 718 mL X min-1 (means +/- SD); 30-s P, 3408 +/- 778 mL X min-1; 4-min ND, 4058 +/- 409 mL X min-1; 4-min P, 3725 +/- 573 mL X min-1). There was no difference between 30-s ND and 4-min ND for peak VO2. The ventilatory anaerobic threshold was not significantly different between any test (30-s ND, 2337 +/- 434 mL O2 X min-1; 30-s P, 2174 +/- 406 mL O2 X min-1; ND, 2433 +/- 685 mL O2 X min-1; 4-min P, 2296 +/- 604 mL O2 X min-1). The VO2 at which blood lactate had increased by 0.5 mM above resting levels was significantly lower than the ventilatory anaerobic threshold for the 4-min ND (1917 +/- 489) and the 4-min P (1978 +/- 412) tests, but was not different for the 30-s ND and 30-s P tests. At exhaustion in the progressive tests, the blood PCO2 was higher (p less than 0.05) in both 30-s tests than 4-min tests.(ABSTRACT TRUNCATED AT 250 WORDS)     

Properties of growth hormone receptors in relation to the adipose conversion of 3T3 cells

Cultured preadipose 3T3 cells undergo a process of differentiation in which they convert to adipose cells. Growth hormone promotes this conversion. Since 3T3 sublines vary in their susceptibility to adipose conversion, it was of interest to examine the properties of the growth hormone receptors in relation to that susceptibility. It was found that preadipose 3T3-F442A cells, which are able to convert to adipose cells with high frequency, are able to bind about 10(4) growth hormone molecules per cell with Kd approximately 10(-9) M. After adipose conversion, no appreciable change in hormone binding was detected. The binding of growth hormone to 3T3-C2 cells (a line virtually insusceptible to adipose conversion) was indistinguishable from that to 3T3-F442A cells. Internalization and degradation of the hormone were also similar in the two cell lines. Susceptibility to adipose conversion is therefore not determined by the relative ability of the cells to bind or degrade the hormone, but must instead depend on some response, as yet unidentified, that follows binding of the hormone.     

The stimulation of glycogenolysis and gluconeogenesis in isolated hepatocytes by opioid peptides.

The opioid agonists [leucine]enkephalin, [D-Ala2,D-Leu5]enkephalin and dynorphin-(1-13)-peptide, but not morphine, stimulated the conversion of [2-14C]pyruvate into glucose and glycogenolysis when added directly to isolated hepatocytes. Naloxone produced a small but significant inhibition of both the basal and stimulated rate of incorporation of label into glucose but had no effect on the total glucose output by the cells. The effects of the opioid peptides were mediated by a cyclic AMP-independent mechanism.     

Transitory hematologic effects of moderate exercise are not influenced by iron supplementation

A young women's exercise/fitness class tested the idea that administration of supplemental iron would prevent "sports anemia" that may develop during exercise and training and improve iron status of exercising females of menstrual age. Fifteen women (aged 18-37) were selected for each of three treatment groups: (1) no supplemental iron; (2) 9 mg X d-1 of Fe; and (3) 18 mg X d-1 of Fe (1 US Recommended Daily Allowance). Women exercised at approximately 85% of maximal heartrate for progressively increasing lengths of time in a jogging program and worked up to 45 min of exercise 4 d X week-1 for 8 weeks. Hematologic analysis was performed in weeks 1, 5, and 8. A significant decline in hemoglobin (Hb) concentration and hematocrit (Hct) was observed at week 5 when all data were examined without regard for iron intake; these red cell indices returned to pre-exercise levels by week 8. Reduction of mean cell hemoglobin concentration (MCHC) indicated that the midpoint decline was not caused by simple hemodilution during exercise. Serum ferritin (SF) concentration changed in parallel with Hb and Hct. Although the midpoint decline in SF was not statistically significant, it ruled out the possibility that turnover of red cell iron was directed to storage. Lowered MCHC and SF suggested lower availability of iron during the synthesis of a new generation of red cells. Few iron treatment effects of magnitude were observed. Iron did not prevent the midpoint decline in Hb concentration. Iron intake did not affect SF, serum iron, transferrin saturation, or final Hb, and Hct.(ABSTRACT TRUNCATED AT 250 WORDS)