Human hemoglobin Portland II (zeta 2 beta 2). Isolation and characterization of Portland hemoglobin components and their constituent globin chains

Two types of embryonic hemoglobins (Hb) containing zeta chains have been identified in the blood of several neonates of Chinese origin with homozygous alpha-thalassemia. In addition to Hb Portland I (zeta 2 gamma 2) which was previously reported, another embryonic hemoglobin has been detected and found to contain zeta chains and beta chains. It is being designated Hb Portland II and has the formula (zeta 2 beta 2). It has a mobility slightly slower than that of Hb A on starch gel electrophoresis at pH 8.6 and has been found in the hemolysates of blood of some but not all hydropic infants. Another component with a mobility faster than that of Hb A2 on starch gel has been isolated from the blood of some hydropic neonates. This latter component is postulated to be zeta 2 delta 2. The occurrence of Hb Portland I and Hb Portland II in these hydropic neonates is consistent with the hypothesis that, in the absence of normal alpha chain production, zeta chains are continued to be produced at later states of development than normal and form tetramers with each of the beta-like globin chains. Because Hb Portland II has not been found in blood from all hydropic neonates, we postulate that the presence of this hemoglobin in these fetuses may be correlated with the gestational age of the fetus at the time of birth.     

Further resolution of the low density lipoprotein spectrum in normal human plasma: physicochemical characteristics of discrete subspecies separated by density gradient ultracentrifugation

The molecular basis of the heterogeneity of plasma low density lipoproteins (LDL, d 1.024-1.050 g/ml) was evaluated in 40 normolipidemic male subjects following fractionation by isopycnic density gradient ultracentrifugation into eight major subspecies. The mass profile of our subjects' LDL uniformly displayed single symmetric or asymmetric peaks as a function of density; the peak occurred most frequently (20 subjects) in subfraction 7 (d 1.0297-1.0327 g/ml). Several physicochemical properties (hydrodynamic behavior, electrophoretic mobility, chemical composition, size and particle heterogeneity, and apolipoprotein heterogeneity) of the LDL subfractions were examined. Hydrodynamic analyses revealed unimodal distributions and distinct peak Sf degree rates in individual subfractions. Such behavior correlated well with particle size and heterogeneity data, in which LDL subspecies were typically resolved as unique narrow bands by gradient gel electrophoresis. Subspecies with average densities of 1.024 to 1.0409 g/ml ranged from 229 to 214 A in particle diameter. LDL protein content increased in parallel with density while the proportion of triglyceride diminished; cholesteryl esters predominated, accounting for approximately 40% or more by weight. Distinct differences in net electric charge were demonstrated by electrophoresis in agarose gel, the subspecies with average density of 1.0314 g/ml displaying the lowest net negative charge. ApoB-100 was the major apoprotein in all subspecies, and constituted the unique protein component over the density interval 1.0271-1.0393 g/ml. ApoE and apo[a] were detected at densities less than 1.0271 and greater than 1.0393 g/ml. While apoE was evenly distributed within these two regions, representing up to 2% of apoLDL, the distribution of apo[a] was skewed towards the denser region, in which it amounted to 3-7% of apoLDL. ApoC-III was detectable as a trace component at densities greater than 1.0358 g/ml. Calculation of the number of molecules of each chemical component per LDL subspecies showed the presence of one copy of apoB-100 per particle, in association with decreasing amounts of cholesteryl ester, free cholesterol, and phospholipid. These data indicate that a similar overall molecular organization and structure is maintained in a unimodal distribution of LDL particle subspecies over the density range approximately 1.02 to 1.05 g/ml. In sum, our data may be interpreted to suggest that microheterogeneity in the physicochemical properties of human LDL subspecies reflects dissimilarities in their origins, intravascular metabolism, tissular fate, and possibly in their atherogenicity.     

Molecular evidences of single mutational events followed by recurrent crossing-overs in the common delta-globin alleles in the Mediterranean area

The human delta-globin gene (HBD) is one of the beta-like globin genes expressed in adults. In the Mediterranean countries the carriers of delta-thalassemia defects or Hb A2-variants are >1% and about 40/70 known alleles have been found in families with this ethnic origin. The scope of this study was to investigate the variability of the gene and of the chromosomal background in order to highlight the origin and spreading of the delta-globin gene alleles in the Mediterranean area. We carried out the characterization of the delta-globin gene alleles and of RFLP-haplotypes, SNPs and one microsatellite associated with them in 231 carriers originating principally from East Sicily. Seventeen alleles were identified, of which five were new. The chromosomes associated with mutated alleles from unrelated carriers were 158; the allele Hb A2-Yialousa accounted for about 75% of relative frequency, Hb A2-Mitsero for about 8%. The alleles were associated with RFLP 5'-haplotypes "- - - -" or "+ - + +", prevalent in the Mediterranean area, except Hb A2-Mitsero associated with the 5'-haplotype "Benin" "- - - +" and the Hb A2' associated with "+ - - +", both of African origin. Each allele showed linkage with one haplotype with these exceptions. The Hb A2-Yialousa showed heterogeneity of the 5'-haplotype in 2/58 chromosomes; the Hb A2-Mitsero showed SNPs and (A)gamma-microsatellite typical of a "Benin" haplotype found associated with the Hb C and Hb S chromosomes; the Hb A2-Yialousa (14/58 chromosomes), Hb A2-Mitsero, Hb A2-Pylos, Hb A2-Fitzroy showed heterogeneity in the 3'-haplotypes and beta-globin gene SNPs. The Hb A2-Coburg was found associated with the haplotype "+ - + +/+ +" different from that already reported "- - - -/+ -". With the exception of this last allele, the linkage of each mutation with a core of RFLPs or SNPs around or inside the delta-globin locus suggested the unicentric origin of the mutations followed by recurrent recombination events causing the chromosomal background heterogeneity.     

Changes in the microheterogeneity of histone H1 after mengovirus infection of Ehrlich ascites tumor cells.

Employing high-resolution polyacryl-amide gradient slab-gel electrophoresis in 6M urea and 6% acetic acid, a distinct change in the microheterogeneity of histone H1 was detected after mengovirus infection of Ehrlich ascites tumor cells. Whereas in uninfected cells the band multiplicity was found to be 6, it was reduced to 4 in the course of the infectious cycle (8 to 9 h). It could be deomonstrated that, while the electrophoretically slowly moving subspecies H1e and H1f disappeared from the band profile of histone H1, the faster migrating bands H1a and H1b increased in relative intensity. The relative intensity of band H1d was also drastically reduced, that of band H1c stayed practically constant throughout infection. When Ehrlich ascites tumor cells were labeled with a mixture of [14C]amino acids prior to mengovirus infection, the radioactivity incorporated into histone H1 subspecies of low electrophoretic mobility was chased into histone H1 components of high mobility in the course of infection, suggesting a virus-induced, unidirectional interconversion of the multiple histone H1 subunits. With the exception of a histone H2B subspecies, which also decreased in amount, the relative quantities of the other histones stayed constant throughout infection.     

Molecular heterogeneity of the subclasses of islet-activating protein (pertussis toxin)-sensitive GTP-binding proteins in porcine thyroid tissue

From porcine thyroid cell membranes, we purified five GTP-binding proteins (G-proteins); Nos. 1 to 3 have 41-kDa alpha-subunits, and Nos. 4 and 5 have 40-kDa alpha-subunits. They were chromatographically (Mono Q) separable and served as specific substrates for islet-activating protein (pertussis toxin). G-proteins 1 and 2 were indistinguishable from porcine brain Gi1 with respect to three criteria, i.e., mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), pI of the ADP-ribosylated alpha-subunit, and immunoreactivity. G-protein 3 was identified as Gi3 by immunoreactivity. The SDS-PAGE and isoelectric focusing (IEF) analyses identified G-proteins 4 and 5 as being chromatographically heterogeneous subtypes of Gi2 in comparison with a pure porcine brain preparation. The IEF analysis also disclosed that each of the Gi1, Gi2, and Gi3 subspecies isolated in the present study has a minor component characterized by a slightly lower pI of its alpha-subunit. We conclude that porcine thyroid tissue contains at least Gi1, Gi2, and Gi3, and that each is made up of heterogeneous populations.     

The hemoglobins of the cold-adapted Antarctic teleost Cygnodraco mawsoni

The blood of the teleost Cygnodraco mawsoni, of the endemic Antarctic family Bathydraconidae, contains a major hemoglobin (Hb 1), accompanied by a minor component (Hb 2, about 5% of total). The two hemoglobins have identical alpha chains and differ by the beta chain. The complete amino acid sequence of the three chains has been elucidated, thus establishing the primary structure of both hemoglobins. The sequences show a 53-65% identity with non-Antarctic poikilotherm fish species; on the other hand, a very high degree of similarity (83-88%) has been found between Hb 1 and the major component of another Antarctic species of a different family. The hemoglobin functional properties relative to oxygen binding have been investigated in intact erythrocytes, 'stripped' hemolysate and purified components of C. mawsoni. The hemoglobins display the Bohr and Root effects, indicating fine regulation of oxygen binding by pH and by the physiological effectors organic phosphates.     

Long-Term Retention of a Relocated Population of Gopher Tortoises

Abstract: Relocations of gopher tortoises (Gopherus polyphemus) in Florida, USA, are frequently employed as mitigation tools when tortoises occupy land desired for development. Here we present information about retention and health of a relocated population of gopher tortoises 17 years after relocation. We combine our 17-year postrelocation data with earlier surveys 1 year and 2 years postrelocation to examine whether retention rates change over time. We also evaluate whether retention rates vary by age and gender. Of 74 gopher tortoises relocated in 1985, 31 were present in 2002. We found a 1-year retention rate of 42%, with retention rates of 100% each year thereafter, when we used the percentage of relocated individuals captured at each survey. We found a 1-year retention rate of 73%, a retention rate of 92% from year 1 to year 2, and an overall retention rate of 98.5% from year 2 through year 17, based on the assumption that all individuals present in later surveys were present in earlier surveys. We found no significant difference in retention rate over the 17-year period for adults and juveniles and for adult males and females. Relocated gopher tortoises showed natural growth patterns, indicating good health, but 35% of these gopher tortoises had ≥1 symptom of upper respiratory tract disease, a disease associated elsewhere with population declines of tortoises. Thus, retention rates of relocated gopher tortoises change over time, with relatively low retention during the first year postrelocation but nearly 100% retention in subsequent years. In general, our study shows that relocations can successfully lead to long-term retention of gopher tortoises, but we predict that without management (e.g., fire management and predator control) this relocated population is not viable.     

云南白族中发现一例Hb J—Lome[β59(E3)Lys→Asn]

作者在云南白族中发现一例快泳异常血红蛋白(Hb)。经化学结构分析,确认这例快泳血红蛋白的β链第59位的赖氨酸被天冬酰胺取代,为Hb J-Lome,此为云南白族中首次发现。     

Discovery of a new HBB haplotype <Emphasis Type="Italic">w2</Emphasis> in a wild-derived house mouse, <Emphasis Type="Italic">Mus musculus</Emphasis>

Genetic characterization of a wild-derived house mouse, Mus musculus, originally collected near Lake Balkhash in the Republic of Kazakhstan, was performed by examining protein polymorphisms and nucleotide sequences for the hemoglobin beta chain (HBB) subunits. Protein electrophoresis, which was performed on a cellulose-acetate plate, showed an independent mobility pattern representing a new, previously undiscovered haplotype. Neighbor-joining analyses of the HBB adult genes, i.e., HBB-T1 and HBB-T2, and the intergenic spacer region showed that the Lake Balkhash mouse possessed genomic components that were mixed from different haplotypes. Compared to the previously determined HBB haplotypes, d, p, and w1, the HBB-T1 gene and ca. 11 kb of the spacer region were most similar to the w1 haplotype; however, the remainder of the spacer region and the HBB-T2 gene were most similar to the d haplotype but may represent a still uncharacterized and divergent haplotype. The recombination event is predicted to have occurred 2.5 kb upstream of the HBB-T2 gene and may have occurred through intersubspecific hybridization between mice of the musculus subspecies group (with the w1 haplotype) and the castaneus subspecies group (with the d-like haplotype). Alternatively, an unknown subspecies group that is equidistantly divergent from each of these subspecies groups may have been involved. Our findings suggest reticulate evolution among the subspecies groups during the evolution of M. musculus.     

ONTOGENY OF CHICKEN HEMOGLOBIN II. CHROMATOGRAPHIC STUDY OF THE HETEROGENEITY OF HEMOGLOBIN IN DEVELOPMENT

Some properties of the carbonmonoxyhemoglobin (HbCO) from chicken embryos of ages 5, 10 and 15 days of incubation, from 1-day posthatching and from adult chickens have been investigated by chromatography on carboxymethylcellulose (CM-cellulose) column and by starch gel electrophoresis.
Chromatogram of the hemoglobin (Hb) from 5-day chicken embryos has shown that it consists of at least 6 components. Starch gel electrophoresis of each isolated component from the column in phosphate (pH 6.8), in borate (pH 8.6) and in formate buffer (pH 1.9) has shown later that there are 3–4 embryonic type Hb components in 5-day embryos.
Chromatogram of the hemoglobin from adult chickens has shown that it consists of at least 4 components, but the examination of each isolated component from the column by electrophoresis in phosphate (pH 6.8), in borate (pH 8.6) and in formate buffer (pH 1.9) has shown that there are 4–6 adult type Hb components in adults.
In ontogenic process, embryonic Hb type is detectable in embryos up to 15 days of incubation. Fetal Hb type, which is not detectable in adult chickens, can be first found in 10-day embryos.     

Plasma lipids and lipoproteins of some members of the order Perissodactyla.

1. The plasma lipoproteins of various members of the order Perissodactyla have been examined by electrophoresis and analytical ultracentrifugation. 2. In the Equidae, high density (alpha) lipoprotein was the major component (80-90%) and low density (beta) lipoprotein (10-20%) the minor component. 3. In the Tapiridae represented by the Malayan tapir (Tapirus indicus), high density and low density lipoproteins were present in approximately equal amounts. 4. In the Rhinocerotidae, the high density lipoprotein characteristic of the Equidae and Tapiridae was absent, and the plasma lipoproteins consisted of a complex group having beta mobility on electrophoresis and a flotation pattern usually associated with low density lipoprotein. 5. The fatty acid composition of plasma lipids was remarkably similar in all members of the Perissodactyla examined, with very high percentages of linoleic acid (greater than 70%) being found in the cholesteryl esters.     

Pregnenolone binding sites in the rat olfactory bulb

High concentrations of pregnenolone and its sulfate have been found in several areas of rat and human brain and seem to be controlled by local mechanisms. In the present experiments we have demonstrated pregnenolone binding sites in the cytosolic fraction of the rat olfactory bulb. The pregnenolone binding component showed a Kd = 2.34 +/- 0.66 x 10(-7) M and Nmax = 7.25 +/- 1.20 pmol/mg protein. Pregnenolone, pregnenolone sulfate and 17OH-pregnenolone competed equally for the binding sites while other steroids were less competitive. Protease and trypsin inhibited binding by 48 and 60% respectively. Sucrose density gradient analysis showed a minor peak at 4.6 s and a major one at 3.6 s. After gel filtration chromatography the pregnenolone binding component appeared as 2 peaks corresponding to molecular weights of approximately 150 and 220 kDa. Heating at 60 degrees C increased binding by 150%. These results indicate that the olfactory bulb pregnenolone binding component is complex in nature. Rat plasma also bound pregnenolone. Plasma binding sites could be partially differentiated from those in the olfactory bulb on the basis of susceptibility to lipoprotein lipase, effect of heating and mobility during polyacrylamide gel electrophoresis.     

Application of shielding boronate affinity chromatography in the study of the glycation pattern of haemoglobin

Human haemoglobin (Hb) may appear in a number of glycated species. The glycation pattern of Hb using shielding boronate affinity chromatography (SBAC) has been studied in the present work. SBAC is a novel separation technique, which eliminates nonspecific boronate-protein interactions by introducing a so-called shielding reagent. Two samples from Bio-Rad (Lyphochek)--one from normal persons' blood with relatively low HbA(1c) level (HbL) and the other from diabetic patients' blood with an elevated HbA(1c) level (HbH)--were used for the investigation. Glycated Hb (GHb) was separated from nonglycated Hb species using Tris as the shielding reagent. Two eluted peaks, eluted peak 1 (E1) and eluted peak 2 (E2), were obtained using a linear gradient elution with Tris. Several bands were observed on isoelectric focusing gel, which showed the same migration positions as Hb adducts, such as HbA(0), which is major Hb component containing two alpha chains and two beta chains; HbA(1c), which is post-translational glycation on the N-terminus of the beta chains of HbA(0); Foetal Hb (HbF), consisting of two alpha chains and two gamma chains; and glutathione Hb (also called HbSSG), which is the result from thiol-disulphide interchain exchange during oxidation of the thiol groups of Hb. In both HbL and HbH samples, E2 exhibited slightly higher amounts of HbF than E1. Electrospray-ionisation mass spectrometry showed that: (1) HbL-E1 was glycated with single glucose on both alpha and beta chains while no observable glycated chains were present in HbL-E2; (2) both HbH-E1 and HbH-E2 were glycated with single glucoses on both alpha and beta chains, however, compared with HbH-E1, HbH-E2 showed a higher relative intensity of the glycated beta chain and lower relative intensity of the glycated alpha chain; and (3) the degree of glycation increased with increasing glycation level of the sample. The amount of HbA(1c) presented in the eluted peaks was further determined using enzymatic digestion of glycated Hb by endoproteinase Glu-C and the subsequent separation and analysis of the digested peptides by reversed-phase high-performance liquid chromatography and capillary electrophoresis. The values of HbA(1c)/HbA(0) of the eluted peaks, i.e. HbL-E1, HbL-E2, HbH-E1 and HbH-E2, were 0.27, 0.19, 0.50 and 0.43, respectively. In both HbL and HbH samples, E1 contained higher amounts of HbA(1c) than E2. This study demonstrates the structural heterogeneity of GHb as well as the possibility of using SBAC to detect glycated species of Hb.     

Characterization of a monoclonal antibody against active pectate lyase from Erwinia carotovora

A monoclonal antibody (2E2) produced against pectate lyase from Erwinia carotovora ssp. carotovora reacted with a 41- and a 44-kilodaltion protein on Western blots of concentrated Erwinia culture supernatants resolved by sodium dodecyl sulfate - polyacrylamide gel electrophoresis. It was unequivocally shown that monoclonal 2E2 reacted with an active form of pectate lyase by affinity purifying the antigen with the monoclonal. The affinity-purified antigen was enzymatically active and moved as a single protein band in a nonequilibrium isoelectric focusing gel. Monoclonal 2E2 reacted with the pectate lyases of a diverse range of E. carotovora ssp. carotovora, ssp. atroseptica, and ssp. betavasculorum strains, as well as with one of three strains of E. chrysanthemi. The electrophoretic mobility of the major protein (44 kilodaltons) that reacted with 2E2 was identical within a subspecies but differed among subspecies.     

HPLC determination of hemoglobins to establish reference values with the aid of statistics and informatics

The purpose of the present study was to establish reference values for hemoglobins (Hb) using HPLC, in samples containing normal Hb (AA), sickle cell trait without alpha-thalassemia (AS), sickle cell trait with alpha-thalassemia (ASH), sickle cell anemia (SS), and Hb SC disease (SC). The blood samples were analyzed by electrophoresis, HPLC and molecular procedures. The Hb A2 mean was 4.30 +/- 0.44% in AS, 4.18 +/- 0.42% in ASH, 3.90 +/- 1.14% in SS, and 4.39 +/- 0.35% in SC. They were similar, but above the normal range. Between the AS and ASH groups, only the amount of Hb S was higher in the AS group. The Hb S mean in the AS group was 38.54 +/- 3.01% and in the ASH it was 36.54 +/- 3.76%. In the qualitative analysis, using FastMap, distinct groups were seen: AA and SS located at opposite extremes, AS and ASH with overlapping values and intermediate distribution, SC between heterozygotes and the SS group. Hb S was confirmed by allele-specific polymerase chain reaction. The Hb values established will be available for use as a reference for the Brazilian population, drawing attention to the increased levels of Hb A2, which should be considered with caution to prevent incorrect diagnoses.     

Analysis of electrophoretic behavior of cytochrome c oxidase subunits. Retardation coefficient versus molecular weight in dodecyl sulfate urea gels.

1. Standard proteins were examined by electrophoresis in highly cross-linked polyacrylamide gels containing sodium dodecyl sulfate and urea. Their behavior was analyzed at a single gel concentration (by molecular weight vs. relative mobility) and at several gel concentrations (molecular weight vs. retardation coefficient). The validity of the latter method of analysis was established for this gel system. 2. Cytochrome c oxidase was subjected to analysis by this method. Compared to standards, subunits I and III showed increased free electrophoretic mobility, while that of subunit V was slightly decreased. The molecular weight values derived were: I, 44 600; II, 22 700; III, 23 500; IV, 16 900; V, 9400; VI, 7600; VII, 4300. The standard errors were all less than +/- 7%. 3. Isolated V and VI were analyzed by two dimensional dodecyl sulfate electrophoresis, in which the second dimension also contained urea. In contrast to their behavior when the holo-enzyme was examined, these isolated subunits V and VI no longer exchange migrating positions relative to each other during the two dimensional analysis. The molecular weight values of the isolated subunits agree with those of the holo-enzyme.     

Heterogeneity in the globin chain composition of a human minor fetal hemoglobin component

The first hemoglobin found to contain an acetyl blocking group was the minor human fetal hemoglobin, Hb FI, present as 10-15% of the total fetal hemoglobin in umbilical cord blood red cells. Acetylation occurs at the amino-terminal glycine of the gamma-globin chain. Assays for the acetyl group by two different methods gave values less than the 2 per tetramer expected for a fully acetylated hemoglobin. We have purified acetylated fetal hemoglobin FIc to homogeneity. The globin chain composition of Hb FIc has been examined by both globin chain separation on CM-cellulose and by tryptic peptide mapping by HPLC. The identities of the gamma globin chains and of the gamma T-1 peptides were confirmed by amino acid analysis. Globin chain separation profiles showed the presence of 22.3 +/- 7.0% of gamma 0 globin (of the total gamma globin) in Hb FIc. Accordingly, the tryptic peptide maps of Hb FIc tetramers also showed the presence of a similar amount of gamma 0T-1 peptide. The gamma 0T-1 peptide was not present in the maps of isolated gamma Ic globin. It is evident that column purified Hb FIc contains a certain percentage of non-acetylated gamma-globin chains, thus indicating a hybrid globin chain composition for this minor fetal hemoglobin component.     

Den Site Selection by the Florida Spotted Skunk

The eastern spotted skunk (Spilogale putorius) is a species of conservation concern in much of its range and has experienced a decline since the early to mid-1990s. But the subspecies that inhabits peninsular Florida, the Florida spotted skunk (S. p. ambarvalis), might still be abundant and is an important nest predator of the endangered Florida grasshopper sparrow (Ammodramus savannarum floridanus). To gain insight on this little-studied subspecies and inform potential management strategies, we conducted a resource selection study on the Florida spotted skunk. We examined 5 hypotheses for den site selection related to den type, vegetation, and landscape characteristics in a dry prairie ecosystem in central Florida. We tracked 36 individual skunks to 757 den sites. Using discrete choice analysis, we found that male and nonbreeding female skunks at our study site were 5 times more likely to select a mammal burrow over a gopher tortoise (Gopherus polyphemus) burrow, that selection of a den site increased 34% for each 1-burrow increase in the number of nearby burrows, and that selection of a den site increased 3% for every 10-cm increase in a visual obstruction index. Similarly, breeding female skunks were more likely to select mammal burrows and shallow depressions over gopher tortoise burrows by 16-fold and 13-fold, respectively, and selection of a den site increased by 75% for every 1-burrow increase in the number of nearby burrows. In contrast to previous studies that occurred in forested, mountainous environments elsewhere in the species' range, our findings suggest that den characteristics might be more important than landscape or vegetation characteristics to Florida spotted skunk den site selection in dry prairie. Additionally, the frequency of prescribed fires on the landscape did not appear to affect Florida spotted skunk den site selection. Thus, Florida spotted skunks in this ecosystem might be landscape generalists, thereby potentially limiting the ability of managers to control nest predation by this subspecies through habitat management. © 2019 The Wildlife Society.     

Geographic genetic architecture of pocket gopher (Thomomys bottae) populations in Baja California, Mexico

Phylogenetic analyses of complete mitochondrial cytochrome b sequences support the monophyly of pocket gopher (Thomomys bottae) populations from the 1000 km length of the Baja California peninsula of Mexico, relative to other geographical segments of the species range in western North America. The Baja California peninsula is an area that encompasses considerable ecomorphological and infraspecific diversity within this pocket gopher species. However, detailed population analyses encompassing 35 localities distributed over the southern half of the peninsula reveal only trivial phylogeographical structure. Rather, most of the 72 unique 500-base pair haplotypes examined from 142 individuals is restricted to single populations, although a few haplotypes are shared broadly across geography. Individual populations are typically comprised of haplotype sets from different branches in a network of relationships. Analysis of molecular variance (amova) indicates that approximately half of the total pool of variation is contained among individuals within local populations, and that only about 25% can be explained by the regional subdivisions of current subspecies distributions or physiographic realms. A hypothesized historical vicariant event that has been causally linked to the phylogeographical structure of other, codistributed species has had little influence on these pocket gopher populations, explaining only 13% of the total variation. The temporal depth, estimated by coalescence parameters, of the haplotype lineage in Baja California is relatively recent, approximately 300,000 generations; both the mismatch distribution of pairwise comparisons and a significantly positive exponential growth estimate support a recent history of expanding populations; but current, or recent past, migration estimates have remained small, are largely unidirectional from north to south, and weak isolation by distance is present. All data suggest that pocket gophers have relatively recently invaded the southern half of peninsular Baja California, with the genetic signature of expansion still evident but with sufficient time having lapsed to result in a weak isolation by distance pattern. The geographical assemblage of sampled populations thus appears as a meta-population, with limited gene flow contrasting with random haplotype loss due to drift in small, localized populations.     

The apparently symmetrical hexagonal bilayer hemoglobin from Lumbricus terrestris has a large dipole moment

The giant approximately 3.6 MDa hexagonal bilayer hemoglobin (HBL Hb) from Lumbricus terrestris consists of 12 213-kDa dodecamers of four globin chains ([b + a + c]3[d]3) tethered to a central scaffold of approximately 36 non-globin, linker subunits L1-L4 (24-32 kDa). Three-dimensional reconstructions obtained by electron cryomicroscopy showed it to have a D6 point-group symmetry, with the two layers rotated approximately 16 degrees relative to each other. Measurement of the dielectric constants of the Hb and the dodecamer over the frequency range 5-100 kHz indicated relaxation frequencies occurring at 20-40 and 300 kHz, respectively, substantially lower than the 700-800 kHz in HbA. The dipole moments calculated using Oncley's equation were 17,300 +/- 2300 D and 1400 D for the Hb and dodecamer, respectively. The approximately threefold higher dipole moment of the dodecamer relative to HbA is consistent with an asymmetric shape in solution suggested by small-angle X-ray scattering. Although a two-term Debye equation and a prolate ellipsoid of revolution model provided a good fit to the experimental dielectric dispersion of the dodecamer, a three-term Debye equation based on an oblate ellipsoid of revolution model was required to fit the asymmetric dielectric dispersion curve of the Hb: the required additional term may represent either an induced dipole moment or a substructure which rotates independently of the main permanent dipole component of the Hb. The D6 point-group symmetry implies that the dipole moments of the dodecamers cancel out. Thus, in addition to a possible contribution from fluctuations of the proton distribution, the large dipole moment of the Hb may be due to an asymmetric distribution of the heterogeneous linker subunits.