ABSTRACT: To elucidate the structure-function relationships in glucose phosphate isomerase (GPI), we established an expression system for human GPI as a fusion protein with glutathione S-transferase (GST) in E. coli. The GST-GPI fusion protein showed affinities for the substrates glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P) similar to those of the native enzyme purified from human red blood cells (RBC). We expressed GPI cDNAs with four distinct disease-causing mutations and examined their enzymatic characteristics. Although each mutation caused reduced thermal stability, an amino acid substitution Thr-5->Ile (T5I) exhibited marked thermal instability, suggesting that the amino-terminal of GPI is important for enzymatic stability. Thr-224 seemed not to be an essential residue, since the amino acid substitution Thr-224->Met (T224M) showed normal substrate affinity in spite of a slight decrease in both specific activity and thermostability. Gln-343 and Asp-539 have been shown to be in close proximity to the putative catalytic sites, and the present study showed that both Gln-343->Arg (Q343R) and Asp-539->Asn (D539N) caused impaired substrate affinity; Q343R showed high Km for both G6P and F6P, whereas D539N showed significantly decreased affinity only for F6P. These results suggest that not only reduced enzymatic stability but also impaired kinetics may disturb RBC metabolism of the GPI variants associated with hereditary hemolytic anemia.

Keywords: Hemolytic anemia, erythroenzymopathy, glycolysis, point mutation, amino acid substitution.

Reprint requests to: Hitoshi Kanno, M.D.,Ph.D., Okinaka Memorial Institute for Medical Research, 2-2-2 Toranomon, Minato-ku, Tokyo 105, Japan, phone: 81-3-3588-1111 Ext. 4415, fax: 81-3-3505-6205, e-mail: hikanno@med.email.ne.jp.

ABSTRACT: It has been suggested that differences in the frequency of the t(14;18) translocation in follicular lymphoma might explain ethno-geographic variation in the incidence of these tumors. We tested Israeli follicular lymphoma patients for the frequency of the t(14;18) translocation, and reviewed the published literature, comparing the frequency in our series with data from different parts of the world.

Tissue specimens from 36 Israeli follicular lymphoma patients were tested for presence of the translocation by PCR amplification of the MBR breakpoint. Twenty-two of the 36 patients (61%) tested positive.

A systematic search of the literature yielded 35 papers reporting the frequency of the t(14;18) translocation in follicular lymphoma. We analyzed cytogenetic data and molecular data separately. For each method, data were pooled from all studies within each of three geographical regions - USA, East Asia and Europe. Pooled data from cytogenetic studies show a low frequency of the translocation in the Far East (38%) compared to the USA (71%), with an intermediate frequency found in Europe (61%). Molecular studies show a similar frequency of the translocation in the Far East and Europe, significantly lower than the frequency in pooled data from American studies. The frequency in our Israeli series is relatively high, comparable to that detected in the USA.

We suggest that the apparent geographical differences we describe are unlikely to be caused by a difference in the biology of the tumor, and are more likely due to technical and methodological factors. We conclude that it is unlikely that differences in the frequency of the t(14;18) translocation explain the difference in the epidemiology of lymphoma between East and West.

Keywords: Follicular lymphoma, epidemiology, t(14;18), Israel, Europe, Asia, United States.

Reprint requests to: Dr. Dina Ben Yehuda, Department of Hematology, Hadassah University Hospital, Kiryat Hadassah POB 12000, Jerusalem, Israel, phone: 672-2-6776744, fax: 672-2-6423067.