Haemoglobins, LX. Primary structure of the major haemoglobin of the sea lamprey Petromyzon marinus (var. Garonne, Loire)

Lampreys belong to the class of Cyclostomata; practically no evolution of these Vertebrates can be noted since Paleozo?c times; lampreys thus appear as a choice material for studying several problems in the field of biochemical evolution. Several monomeric haemoglobins can be characterized in the erythrocytes of the sea lamprey (Petromyzon marinus). The major constituent was isolated by chromatography, and submitted to tryptic digestion; soluble tryptic peptides were separated by gel filtration into 5 fractions; the peptides of each fraction were isolated either by Dowex-50 chromatography or by HPLC; the insoluble core was oxidized and submitted to HPLC fractionation. The primary structure of the whole chain and of the purified tryptic peptides was determined using automatic sequencing; alignment of the peptides was achieved by homology with the previously established covalent structure of the globin of Lampetra fluviatilis. The sequence we established confirms the crystallographic data of Hendrickson and Love. Globin/haem contacts are discussed; a tentative explanation of the absence of tetramerization can be proposed after comparison with the aminoacid residues involved in alpha 1 beta 1 and alpha 1 beta 2 contacts. Petromyzon globin differs at three locations (Thr/Ser3, Leu/Met58, Thr/Ser60) from Lampetra fluviatilis globin. The monomeric chain of another Cyclostomata Myxine glutinosa, differs more considerably (88 residues). Our results corroborate recent paleontologic data which favour the separation of lampreys from hagfishes; Cyclostomata cannot be considered as a monophylic group. Finally, there is a closer relation between lamprey globin and alpha chains than between this monomeric globin and beta chains, and furthermore apomyoglobins of higher vertebrates.     

Differences in the clotting of lamprey fibrinogen by lamprey and bovine thrombins

1. Improved methods for the purification of lamprey thrombin and fibrinogen are presented. 2. Lamprey thrombin releases two fibrinopeptides from lamprey fibrinogen during the transformation into fibrin. Bovine thrombin releases only one of these, a peptide referred to as fibrinopeptide B. The differences in the by-products of fibrin formation are reflected in the different N-terminal amino acid compositions of the two types of fibrin. 3. The fibrinopeptide that is not removed from the lamprey fibrinogen by bovine thrombin can subsequently be released by treatment of that fibrin with lamprey thrombin. 4. Under the conditions used, lamprey thrombin releases both fibrinopeptides at about the same rate. 5. The differences in interaction among these pairs of related proteins are extreme manifestations of the phenomenon loosely referred to as `species specificity'.     

Characterization of lamprey fibrinopeptides

1. Lamprey fibrinopeptide B is a relatively large peptide made up of about 40 amino acid residues. The peptide is highly electronegative, containing a large number of aspartic acid residues and a tyrosine O-sulphate residue. 2. The amino acid sequence of the first 18 residues from the N-terminal end of fibrinopeptide B has been established. The C-terminal ends with the sequence Val-Arg. Fibrino-peptide B is released by both lamprey and bovine thrombins. 3. Lamprey fibrino-peptide A is a short peptide containing only eight residues. The proposed amino acid sequence is: Asp-Asp-Ser-Ile/Leu-Asp-Ser-Leu/Ile-ArgThis peptide is released by lamprey thrombin but not by bovine thrombin.     

Primitive haemoglobin

Summary The sequences ofPetromyzon andAplysia globins are compared with the postulated vertebrate and mollusc-vertebrate ancestors to see if differences exist in the rates of evolution of different types of residue positions. Between the mollusc-vertebrate ancestor andAplysia globin there is no very striking pattern of changes except that the interior positions are relatively conserved. In the evolution ofPetromyzon haemoglobin, the _{1 2} contact area is relatively conserved. The homopolymeric binding of lamprey Hb seems to be a primitive function.     

Functional properties of lamprey electroreceptors

Spike discharges in nerve fibers, evoked by stimulation of electroreceptors by an electric field directed along the body axis (square pulses of current and a sinusoidal current) were recorded in the lampreyLamperta fluviatilis (L.). Excitation of electroreceptors was shown to arise through the action of the cathode. Minimal values of electric field at which appreciable changes took place in spike activity were 10–60 µV/cm for different nerve fibers. The optimal frequency range of sinusoidal electrical stimulation was 0.05–0.5 Hz. It is suggested that electroreceptors of the Agnatha (caudata) and of gnathostomatous cartilaginous fish share a common origin.Murmansk Marine Biological Institute, Kola Branch, Academy of Sciences of the USSR, Dal'nie Zelentsy, Murmansk Region. Translated from Neirofizioloigya, Vol. 16, No. 1, pp. 105–110, January–February, 1984.     

Fibril-forming collagens in lamprey

Five types of collagen with triple-helical regions approximately 300 nm in length were found in lamprey tissues which show characteristic D-periodic collagen fibrils. These collagens are members of the fibril forming family of this primitive vertebrate. Lamprey collagens were characterized with respect to solubility, mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, carboxylmethyl-cellulose chromatography, peptide digestion patterns, composition, susceptibility to vertebrate collagenase, thermal stability, and segment long spacing-banding pattern. Comparison with fibril-forming collagens in higher vertebrates (types I, II, III, V, and XI) identified three lamprey collagens as types II, V, and XI. Both lamprey dermis and major body wall collagens had properties similar to type I but not the typical heterotrimer composition. Dermis molecules had only alpha 1(I)-like chains, while body wall molecules had alpha 2(I)-like chains combined with chains resembling lamprey type II. Neither collagen exhibited the interchain disulfide linkages or solubility properties of type III. The conservation of fibril organization in type II/type XI tissues in contrast to the major developments in type I and type III tissues after the divergence of lamprey and higher vertebrates is consistent with these results. The presence of type II and type I-like molecules as major collagens and types V and XI as minor collagens in the lamprey, and the differential susceptibility of these molecules to vertebrate collagenase is analogous to the findings in higher vertebrates.     

Type II collagen of lamprey

The major collagen in lamprey notochord is type II, as determined by its amino acid composition and solubility properties. This collagen has a distribution of charged residues indistinguishable from higher vertebrate Type II collagens as judged by its SLS banding pattern. Lamprey type II collagen has a higher thermal stability than lamprey skin collagen, in contrast to the identical melting temperatures for these types in mammals. A minor collagen in lamprey notochord has solubility properties, amino acid composition, and electrophoretic mobility similar to that of 1 alpha, 2 alpha, 3 alpha collagen in human cartilage.     

Effect of ionizing radiation on haemoglobin: the oxy-derivative of haemoglobin Iwate

It is well established that exposure of oxyhaemoglobin to ionizing radiation results in remarkably selective electron addition to the (FeO2) unit, giving a novel species, (FeO2)-, in which the extra electron is largely localized on iron and dioxygen. This work has now been extended to haemoglobin (Hb.) Iwate. The haemoglobin M. Iwate used is a mutant haemoglobin having only Fe(III) alpha-chains by oxy beta-chains (alpha 2 Met beta 2 oxy). The haem iron atoms in the alpha-chains are coordinated in the fifth site by a proximal tyrosine in place of histidine. This unit is a high-spin Fe(III) with axial symmetry and prominent electron spin resonance (ESR) features in the g = 6 and g = 2 regions. On exposure to 60Co gamma-rays at 77 K, efficient electron addition occurred at both types of iron centre, giving Fe(II) and (FeO2)- units. The former was monitored by the decrease of the g = 6 feature for Fe(III) and the latter by the growth of g-features at 2.254 (gx), 2.149 (gy) and 1.967 (gz). These values are close to those for the FeO2- centre formed in the beta-chains of normal oxyhaemoglobin. On annealing above 77 K, two changes occurred: first there was a small but clear increase in gx and gy, followed by a marked reduction in gx and gy giving g-values close to those for the centre formed directly in the alpha-chains of the normal protein. Finally, this intermediate species gave a centre having gx = 2.310, gy = 2.180 and gz = 1.935. These values are typical of low-spin Fe(III) haemoglobin and are assigned to the protonated complex, Fe(III)O2H. Ultimately at ca. room temperature, this was converted into the high-spin, met-form, with a gain in the g = 6 feature. These results established that the beta-chain centre in Hb. Iwate behave in the same way as isolated beta-chains. They also confirm that electron addition to the oxy-units is facile, even in the presence of Fe(III) units in each tetramer. The results also confirm that electron capture to give (FeO2)- units is not followed by internal electron-transfer to give Fe(II) from the Fe(III) centres in the alpha-chains.     

Reproductive ecology of the river lamprey

The reproductive ecology of river lamprey Lampetra fluviatilis was investigated during the spawning period 2003 in the River Derwent, north‐east England. Over this period 1199 ± 104 individuals day⁻¹(mean ±  s . d .) were counted on one spawning site (area c . 450 m²), but mark‐recapture estimates suggested that >5000 river lamprey used this site over the same period and egg deposition was estimated as 168 000 eggs m⁻². The operational sex ratio of river lamprey in spawning clusters changed between spawning phases, from domination by females during the nest‐building phase (male : female ratio, 1 : 3·46), to a preponderance of males during the spawning phase (male : female, 1 : 0·37), followed by a return to a majority of females after spawning (male : female, 1 : 3·74). Recapture data showed that >97% of recaptured, tagged males were recorded at two or more nests, whereas almost 50% of recaptured, tagged females were recorded at the same nest, suggesting a promiscuous mating system, with a tendency towards polygyny within the population. Within the lower 80 km of the River Derwent and its tributaries, evidence of river lamprey spawning was found at only six sites, and most spawning (>80% of the observed spawning population) was at one site.     

The lamprey in evolutionary studies

Lampreys are a key species to study the evolution of morphological characters at the dawn of Craniates and throughout the evolution of the craniate’s phylum. Here, we review a number of research fields where studies on lampreys have recently brought significant and fundamental insights on the timing and mechanisms of evolution, on the amazing diversification of morphology and on the emergence of novelties among Craniates. We report recent example studies on neural crest, muscle and the acquisition of jaws, where important technical advancements in lamprey developmental biology have been made (morpholino injections, protein-soaked bead applications or even the first transgenesis trials). We describe progress in the understanding and knowledge about lamprey anatomy and physiology (skeleton, immune system and buccal secretion), ecology (life cycle, embryology), phylogeny (genome duplications, monophyly of cyclostomes), paleontology, embryonic development and the beginnings of lamprey genomics. Finally, in a special focus on the nervous system, we describe how changes in signaling, neurogenesis or neuronal migration patterns during brain development may be at the origin of some important differences observed between lamprey and gnathostome brains.