IDENTIFICATION OF THE CDNA FOR HUMAN RED BLOOD CELL-SPECIFIC HEXOKINASE ISOZYME

A unique cDNA for hexokinase (HK) was identified from poly(A)(+) RNA o f human reticulocytes by anchored polymerase chain reaction. This appe ared to represent the cDNA for the red brood cell (RBC)-specific HK is ozyme (HKR) described in our previous study (Murakami et al: Blood 75: 770, 1990). Its nucleotide sequence was identical to HKI cDNA except f or the 5' extreme end. It lacked the first 62 nucleotides of the HKI c oding region: instead, it contained a unique sequence of 60 nucleotide s at the beginning of the coding sequence as well as another unique se quence upstream of the putative translation initiation site. It lacked the porin-binding domain which facilitates binding to the mitochondri a, thus explaining the exclusive cytoplasmic localization of HKR. It w as the major cDNA derived from reticutocytes, consistent with the obse rvation that HKR activity is predominant in reticulocytes. Northern bl ot analysis showed that it was expressed in the reticulocytes and in t he K562 erythroleukemic cell line, but not in a lymphocytic cell line. In the extract of K562 cells, HKR activity co-eluted with the HKR of human RBCs on a MonoQ column (Pharmacia, Piscataway, NJ) chromatograph y, using a salt gradient elution. The separate genetic control of the RBC-specific HK isozyme explains the clinical reports of two types of HK deficiency, one in which the HK activity was reduced exclusively in the RBC (HKR defect) and another with general decrease of HK activity in several tissues (HKI defect). (C) 1997 by The American Society of Hematology.