Michael Zasloff

A two-dimensional NMR study of the antimicrobial peptide magainin 2

Using two‐dimensional NMR spectroscopy, a complete 1H resonance assignment has been obtained for the peptide magaining 2 recently isolated from Xenopus laevis. It is demonstrated that this peptide adopts an α‐helical structure with amphiphilic character when dissolved in a mixture of trifluoroethanol (TFE) and H2O. The transition to the α‐helical conformation occurs at very low concentrations of TFE.

Antimicrobial properties of peptides from Xenopus granular gland secretions

Previously, we described a family of novel broad spectrum antimicrobial peptides, magainins, from the skin of Xenopus laevis. In this report we show that at least two other Xenopus peptides, present in the skin and its secretions, PGLa and a peptide released from the xenopsin precursor, exhibit antimicrobial properties comparable to the magainins. The identification of these newer members provides insight into the structural diversity of vertebrate antimicrobial peptides.

Magainin 2, a natural antibiotic from frog skin, forms ion channels in lipid bilayer membranes

We have examined the ion channel forming properties of magainin 2 by incorporating the peptide into artificial lipid bilayers held under voltage clamp. Magainin 2 increased lipid bilayer conductance in a concentration dependent manner with a Hill coefficient of 1.7. The magainin 2 conductance was selective for monovalent cations over anions with a ratio of 5:1 and had both voltage-sensitive and -insensitive components. Two structurally related but antibiotically less potent analogues, magainin 1 and Z-12, also increased lipid bilayer conductance with a similar ion selectivity but these peptides were less potent than magainin 2. We propose that the weak cation selectivity of the magainin channels can be accounted for by the inclusion of negatively charged lipids in the channel complex and suggest two possible structures for such a channel. The ionophoric properties of these peptides are likely to be proximal to their antibiotic activities.

Comparative analysis of human chromosomal segments bearing nonallelic dispersed tRNAimet genes

About 12 tRNAimet genes have been found at scattered locations in the human genome. Four fragments of human fetal liver DNA ranging in size from 11 to 18 kb, each containing a single tRNAimet gene, were cloned from a recombinant phage library. On the basis of restriction site mapping, electron microscopic analysis of heteroduplex structures and the maps of sequences transcribed in vivo in human fibroblasts, obtained by a novel contract-hybridization method, the fragments were shown to represent two different loci with homologies limited to several dispersed repetitive sequences within each of the chromosomal neighborhoods. Detailed structural analysis of the tRNA regions revealed several blocks of homology at the proximal flanking sequences of the two nonallelic genes, one of which differed from the common vertebrate tRNAimet sequence by a base substitution at position 56 with a T in place of G. Two oligonucleotides identical or very similar to tRNAimet structural sequences were present at the 5 border of both genes. A review of published sequence data showed other unequivocal examples of tRNA-coding sequences present at the 5 flanking region of the associated tRNA gene, which in several cases contained the site of transcription initiation.

Electroretinographic Findings in the Mucopolysaccharidoses

The degree of retinal involvement of 20 patients (ages 3-21) with mucopolysaccharidosis (MPS) types I, II, and III was assessed using electroretinography (ERG) under standardized conditions. ERG evidence of retinal dysfunction ranged from none to severe in MPS I and II, and from moderate to severe in MPS III. The ERG abnormalities were common to all three types of MPS, and showed the pattern seen in rod-cone degenerations, where the rod-mediated responses are more severely affected than the cone-mediated responses. The ophthalmoscopic signs were less striking than the electrophysiologic findings, and were usually restricted to mild changes of the retinal pigment epithelium.

Failure of surgery and isotretinoin to relieve jaw immobilization in fibrodysplasia ossificans progressiva: Report of two cases

Two patients with fibrodysplasia ossificans progressiva who presented with jaw immobilization due to formation of bone between the maxilla and mandible were treated with surgical resection of their ectopic bone in conjunction with experimental, adjunctive medical therapy using isotretinoin. Both patients had recurrence of their ectopic ossification within 2 months of surgery. Surgery to remove joint-bridging ossifications in FOP is not recommended.

Enhancement of mRNA nuclear transport by promoter elements

This report describes studies on the kinetics of herpes thymidine kinase (TK) mRNA transport in X. laevis oocytes as studied by microinjection and microdissection. We show that TK mRNA nuclear transport in this cell can be dramatically altered by introduction of specific DNA sequences into the nucleus. Introduction of DNA sequences encompassing the promoter results in enhancement, in trans, of a transport process that can deliver previously synthesized RNA to the cytoplasm. The report suggests that the promoter of a eukaryotic gene can functionally interact with the mechanism involved in mRNA nuclear transport.

Technetium-99m MDP demonstration of heterotopic ossification in fibrodysplasia ossificans progressiva.

The extensive heterotopic bone formation in patients with fibrodysplasia ossificans progressiva (FOP) has been documented previously with radiographs. A case in which a Tc-99m MDP bone scan showed increased uptake at sites well before ossification could be documented radiographically is described. This finding suggests that bone scans would likely be useful to monitor the extent of involvement with FOP and to detect areas of new activity.

843 AMNIOTIC MEMBRANE IMPLANTATION IN MUCOPOLYSACCHARIDOSIS

Amniotic membrane implantation has been performed as an approach to enzyme replacement in mucopolysaccharidosis (MPS). The rationale for this study is based on the findings that cultured amniocytes secrete lysosomal enzymes and that amnion membranes are non-immunogenic. 19 patients (MPS I,II,III), ages 3 to 16 yr., have had implantation of human amniotic membranes obtained from elective repeat C-sections. The subcutaneous implants in the abdominal wall have been well tolerated with follow-up between 4 to 12 mo. The effects of amniotic membrane implantation on the clinical status of the patients was evaluated by comparing the following studies pre- and 6 mo. post-implantation: EKG, echocardiography, head and liver CT, liver/spleen scan, skeletal survey, fundus exam, electroretinography, audiological assessment, auditory brain stem response and joint range of motion. Objective clinical improvement was noted as an increased range of motion in 3 of 18 patients. Biochemical analysis demonstrated no change in serum or WBC enzyme activity. Urinary GAG transiently decreased in 2 of 10 patients with levels returning to pre-treatment range at 6 mo. post implantation. Biopsy of the implantation site at 6 mo. post implantation demonstrated a foreign body reaction with no evidence of amnion tissue. Amniotic membrane implantation appears to have limited success as a means of enzyme replacement.