Extensive homology between the subunits of the phytohemagglutinin mitogenic proteins derived from Phaseolus vulgaris.

Abstract

The phytohemagglutinin mitogenic proteins derived from Phaseolus vulgaris comprise a class of five glycoproteins that are isomeric tetramers composed of varying proportions of two different subunits (L and R). Within the native tetramer, the L subunit is a potent leukoagglutinin and mitogen that lacks hemagglutinating properties, whereas the R subunit is a potent hemagglutinin with little or no mitogenic activity. The subunits have been isolated in homogeneous form by isoelectric focusing in 8 M urea. Previous work has shown that they have equal molecular weights and differ in amino-acid sequence from residues 1-7, but are identical in positions 8-24 [(1973) J. Exp. Med. 138, 939-9511. We now report amino-acid composition studies which reveal striking similarities between the subunits. Both lack methionine and cysteine. The twelfth residue in each subunit is a glycosylated asparagine, with the identical carbohydrate composition in each. The last three residues of the subunits, as determined by carboxypeptidase A digestion, are identical. Tryptic peptide mapping of the succinylated phytohemagglutinin subunits reveals a high degree of similarity. We conclude that the substantial difference in biological properties among the tetrameric phytohemagglutinin mitogens is a result of relatively restricted differences in the primary structure of their constituent subunits.The phytohemagglutinin (PHAP) mitogenic proteins derived from the red kidney bean, Phaseolus vulgaris, have been shown to comprise a family of five heterogeneous proteins (1, 2). They consist of isomeric, noncovalently bound tetramers which are made up of two different subunits, designated as L and R ( Fig. 1) (2-4). One of the five proteins is a potent leukoagglutinin with low hemagglutinating activity (L-PHAP); it is homogeneous, consisting of four identical subunits (1). Three other, closely related proteins, which have modest leukoagglutinating but potent hemagglutinating properties (H-PHAP), have also been isolated. They consist of hybrid tetramers containing varying proportions of the two subunits (2-2R, 1L3R, and 4R), the increasing R subunit content of which is reflected by increasingly cathodal migration on polyacrylamide gel electrophoresis. A fifth PHAP mitogenic protein (3L-1R) has been identified, but detailed study of its properties has been hampered in the past by its contamination with other proteins. We have tentatively concluded that, within each native tetramer, the L subunit has strong mitogenic activity and a high affinity for receptors of lymphocyte membranes, but little or no affinity for those of erythrocytes.Conversely, the R subunit has a high affinity for erythrocyte membrane receptors, but little for those of lymphocytes. As a result, the 4R tetramer displays little or no lymphocyte mitogenic activity (6). The hybrid molecules (1L-3R, 2L-2R, and 3L-1R tetramers) have been found to be mitogenic, to cause mixed agglutination of erythrocytes and lymphocytes, and to have enhanced lymphocyte-transforming capability in the p...

Isolation of two blood type A and N specific isolectins from Moluccella laevis seeds

“…Furthermore, we cannot conclude whether the isolectins consist of subunits which are anti-A + N, or whether they are a mixture of molecular species made up of varying proportions of anti-A and anti-N subunits. The latter case would be analogous to what has been observed with other lectins, such as PHA [14], Griffonia simplicifolia lectin I [15] and Maclura pomifera lectin [16].…”

cDNA cloning and in vitro synthesis of the Dolichos biflorus seed lectin

“…In the case of phytohemagglutinin, the stoichiometric assembly of the two different subunit types does not appear to be strictly controlled during lectin synthesis. This results in a heterogeneous mixture of mature lectin proteins consisting of the five possible combinations of the E and L subunits into the tetramer [39]. The D. bijlorus seed lectin is consistently isolated as a tetramer with a 2:2 stoichiometric configuration of subunit I and subunit 11.…”

Mitogenic leukoagglutinin from Phaseolus vulgaris binds to a pentasaccharide unit in N-acetyllactosamine-type glycoprotein glycans.

“…Each isolectin is a tetramer OfMr 115,000 (3) in which the subunits are held together by noncovalent forces (1)(2)(3)(4). The subunits are of two different types, designated leukocyte reactive (L) and erythrocyte reactive (E), and occur in the combinations L4, L3E, L2E2, LE3, and E4 (1)(2)(3). L has high affinity for lymphocyte surface receptors (1)(2)(3)(4)(5) but little for those of erythrocytes (refs.…”

“…The subunits are of two different types, designated leukocyte reactive (L) and erythrocyte reactive (E), and occur in the combinations L4, L3E, L2E2, LE3, and E4 (1)(2)(3). L has high affinity for lymphocyte surface receptors (1)(2)(3)(4)(5) but little for those of erythrocytes (refs. 1 and 2; unpublished data) and is responsible for the mitogenic properties of the isolectins (1)(2)(3)(4)(5).…”

“…Red kidney bean (Phaseolus vulgaris) extract contains a family offive isolectins (1)(2)(3). Each isolectin is a tetramer OfMr 115,000 (3) in which the subunits are held together by noncovalent forces (1)(2)(3)(4).…”