Kiso Akahane

Possible Link between Glycolysis and Apoptosis Induced by Sodium Fluoride

Fluoride has been used to prevent caries in the dentition, but the possible underlying mechanisms of cytotoxicity induction by this compound are still unclear. Since fluoride is known as an inhibitor of glycolytic enzymes, we investigated the possible connection between NaF-induced apoptosis and glycolysis in human promyelocytic leukemia HL-60 cells. NaF-induced apoptotic cell death is characterized by caspase activation, internucleosomal DNA fragmentation, loss of mitochondrial membrane potential, and production of apoptotic bodies. Higher activation of caspases-3 and -9, as compared with that of caspase-8, suggested the involvement of an extrinsic pathway. Utilization of glucose was nearly halted by NaF, whereas that of glutamine was rather enhanced. NaF enhanced the expression of Bad protein, but not that of Bcl-2 and Bax proteins, and reduced HIF-1α mRNA expression. Analysis of these data suggests a possible link between glycolysis and apoptosis.

Recovery of tryptophan from 25-minute acid hydrolysates of protein

It was found that thioglycolic acid prevents destruction of tryptophan during rapid hydrolysis of protein with a trifluoroacetic acid/HCl mixture (1:2, v/v) at 166 degrees C for 25 or 50 min. The addition of 5% (v/v) thioglycolic acid gave the maximum tryptophan recovery (88.3%) for a 25-min hydrolysate of lysozyme. Tryptophan recoveries varied slightly among three different proteins; 88% for lysozyme, 73% for alpha-chymotrypsinogen A, and 85% for apomyoglobin. However, when extrapolated to zero time, the values were close to one another: 94, 87, and 88%, respectively. The addition of thioglycolic acid was also advantageous for recovering amino acids other than tryptophan. Particularly, yields of carboxymethylcysteine and methionine were greatly improved. This modified rapid hydrolysis method gave satisfactory results without the need for separate analyses of tryptophan and cysteine, provided proteins were reduced and carboxymethylated prior to hydrolysis.

The N-terminal sequence of rat pepsinogen

The amino-acid sequence of 96 residues in the N-terminal region of rat pepsinogen I was determined and the first 46 residues were found to constitute the activation peptide segment. There was high degree of homology between the activation segments of rat pepsinogen and some pepsinogens A (pig, cow, Japanese monkey and human). However, the number of residues substituted between rat and the other pepsinogens were considerably larger than those among pepsinogens A. In the N-terminal 24 residues of active pepsin, homology (88%) between rat pepsin and human gastricsin was higher than that (50%) between rat pepsin and pepsin A from human or pig. This strongly suggests that rat pepsin should be classified as pepsin C.

Nucleotide Sequences of Pigeon Feather Keratin Genes

We analyzed two pigeon feather keratin clones from a cosmid pigeon genomic library. Each of the clones contained three feather keratin genes that had the same general structure: a 5′ non-coding region separated by an intron, a protein-coding region encoding a protein of 100 amino acids, and a 3′ non-coding region. Length and transcriptional organization of the genes were variable. The length variation, about 1.2–3.7 kb, was mainly due to the difference in the length of the 3′ non-coding region, and the longer genes had opposite transcriptional organization in contrast to the shorter genes. The nucleotide sequences of the coding region were very similar among the six genes but not the same.