Recognition of specific Physarum alpha-tubulin isotypes by a monoclonal antibody. Sequence heterogeneity around the acetylation site at lysine 40

The monoclonal antibody 6-11B-1 recognises specifically the acetylated form of alpha-tubulin. The acetylation event occurs on a unique lysine residue, lysine 40. Using 6-11B-1, acetylated alpha-tubulin was detected in myxamoebae but not plasmodia of Physarum polycephalum. Following chemical acetylation plasmodial alpha-tubulin was detected by 6-11B-1. The monoclonal antibody KMP-1 recognises certain Physarum alpha-tubulin isotypes but only in non-acetylated form. Whilst recognising all the non-acetylated fraction of myxamoebal alpha-tubulin only a proportion of plasmodial alpha-tubulin was recognised by KMP-1. Peptides were synthesised corresponding to the acetylation domains (containing lysine 40) of myxamoebal alpha-tubulin and the inferred acetylation domains of two plasmodial-specific alpha-tubulin isotypes. The only difference between the two peptides was at a single residue corresponding to amino acid 44 in the polypeptide. Tyrosine was present in myxamoebal alpha-tubulin and glycine was present in the plasmodial specific peptides; the peptides are referred to as the Tyr44 and Gly44 peptides respectively. Both peptides in acetylated form blocked 6-11B-1 reactivity towards acetylated myxamoebal alpha-tubulin. The Tyr44 but not the Gly44 peptide blocked KMP-1 reactivity towards non-acetylated myxamoebal alpha-tubulin. Tyrosine at position 44 is not found in any other known alpha-tubulin. Thus a unique antigenic determinant exists in certain Physarum alpha-tubulin isotypes, close to the acetylation site at lysine 40. This antigenic determinant forms part of the KMP-1 recognition epitope and explains the unique isotype selectivity of this monoclonal antibody.     

ATP-dependent peptide release from mitochondria of reticulocytes

ATP-dependent release of TCA-precipitable peptides from mitochondria-containing stroma (MCS) is described. The process is independent of ubiquitin, but is sensitive to hemin and to heat treatment. Neither chloramphenicol nor EGTA inhibit. 50% of the activity is dependent on charged tRNA. The peptides released from MCS possess a molecular mass of about 1–5 kDa and are degraded to TCA-soluble compounds by a cytosolic protease system (fraction II) without ubiquitin.     

Malaria vaccines--progress and problems.

Developing a vaccine against malaria is a major priority of the WHO. A decade of research exploiting the techniques of molecular biology has yielded a series of potentially protective vaccines. However, progress has been frustrated by the complexity of the parasite's life cycle and the antigenic diversity exhibited by each stage. Although candidate vaccines are now entering human trials, questions still arise concerning the nature of a successful malaria vaccine and who will benefit from it.     

Computer stimulation of sedimentation in the ultracentrifuge. VII. Solutes undergoing indefinite self-association

The velocity sedimentation of solutes involved in self-association equilibria of the indefinite type was simulated using a computer model. The changes in boundary shape that resulted from varying the association constant and the molecular weight of the self-associating monomer were examined. Both ideally and nonideally sedimenting solutes were considered, and several alternative treatments of the variation of the frictional ratio with molecular size were used. All of the calculated boundaries were skewed, with the leading limb of the gradient profile steeper than the trailing limb. For relatively tightly self-associating solutes, the boundaries were very broad and strongly skewed. No shoulders or subsidiary peaks were observed for any of the model solutes used.     

Antiviral agents 2. Synthesis of trimeric naphthoquinone analogues of conocurvone and their antiviral evaluation against HIV

The synthesis of a new series of conocurvone analogues is presented that explores the importance of the pyran rings of conocurvone, their degree of unsaturation as well as the role of alkoxy functionalities as pyran ring replacements, for the inhibition of the HIV-1 integrase (IN) enzyme. Difficulties in synthesising a trimeric naphthoquinone where the central quinone bears a peri-dihydropyran ring was attributed to distortion of the electrophilic dihaloquinone successfully utilised in the past. Increased electron density could also be a factor in reducing reactivity. The desired central dihydropyran bearing trimeric naphthoquinone was successfully synthesised by using a more reactive bromo-tosyloxyquinone intermediate. A maleimide derivative, where the central quinone between the pendant hydroxyquinones was replaced, was successfully synthesised and although it exhibited comparable enzyme inhibitory activity it had negligible HIV inhibitory cellular activity. Compounds were assessed for activity in both in vitro assays using purified recombinant HIV-1 IN and demonstrated superior or comparable activity to conocurvone derivatives previously reported.