Insect Biochem. Molec. Biol. Vol. 27, No. 7, pp. 663–670, 1997  1997 Elsevier Science Ltd Pergamon PII: S0965-1748(97)00042-8 All rights reserved. Printed in Great Britain 0965-1748/97 $17.00 + 0.00 Lipophorin: A Hemolymph Juvenile Hormone Binding Protein in the German Cockroach, Blattella germanica VEERESH L. SEVALA,² JANE A. S. BACHMANN,² COBY SCHAL²* Received 14 January 1997; revised and accepted 4 June 1997 We examined the binding of [ 3 H](10R) juvenile hormone (JH) III to lipophorin that was pur- ified from the hemolymph of Blattella germanica. Binding was found to be specific, saturable and with high affinity to JH III. Using Scatchard analysis, the equilibrium dissociation con- stant (K d ) and total binding capacity (B max ) were estimated to be 9.75 ± 0.64 nM and 0.241 ± 0.02 nmol/mg protein, respectively. Competitive displacement studies with racemic JH III, JH I, cuticular hydrocarbon, contact sex pheromone, and the JH analogs pyriproxyfen, fenoxy- carb, and hydroprene showed that only JH III readily displaced [ 3 H](10R)JH III from the binding site. However, hydroprene competed for the JH III binding site more effectively than the other two JH analogs. Photoaffinity labelling using the JH III analog [ 3 H]epoxyfarnesyl diazoacetate demonstrated that the JH binding site was on apolipophorin-I, the large subunit of the lipophorin complex.  1997 Elsevier Science Ltd Blattella germanica Lipophorin Juvenile hormone Binding protein Hemolymph Pyriproxyfen Fenoxycarb Pheromone Photoactivation INTRODUCTION Juvenile hormones (JH) play important roles in insect metamorphosis and reproduction. Juvenile hormones are hydrophobic (solubility limit of 54 M in aqueous buffer; Trowell, 1992), and it has been well established from various studies that JH is transported from the corpora allata (CA) to target tissues by plasma JH bind- ing proteins (JHBP) (Goodman, 1990; Trowell, 1992). In Manduca sexta virtually all JH in hemolymph exists as a complex with JHBP (Hidayat and Goodman, 1994). Whitmore and Gilbert (1972) first reported the occur- rence of a JHBP in the hemolymph of Hyalophora glov- eri. Since then, several such proteins have been isolated and characterized from a number of insect taxa (Goodman and Chang, 1985; Trowell, 1992; Prestwich et al., 1994). On the basis of their molecular weight and affinity to JH, three different types of JHBPs have been identified. In M. sexta and other Lepidoptera, low mol- ecular weight JHBPs (M r  30 kDa) have been shown to have relatively high affinity (dissociation constant 72– *Author for correspondence. Tel: + 1 919 515 1821; Fax: + 1 919 515 7746; E-mail: coby schal@ncsu.edu. ²Department of Entomology, North Carolina State University, Raleigh, NC 27695-7613, U.S.A. 663 650 nM) to JH I and JH II (for review see Goodman and Chang, 1985; Trowell, 1992). The other two types are high molecular weight lipoproteins. One is a very high density lipoprotein (d = 1.25 mg/ml) that functions as a JH III transporter in Locusta migratoria; it is composed of six identical subunits (77 kDa), each of which contains a JH binding site (Koopmanschap and de Kort, 1988). The other is lipophorin (Lp), a high density lipoprotein which has a single site that binds JH III with high speci- ficity and high affinity (K d for racemic JH III = 1.5 to 157 nM). This type of JHBP has been isolated from sev- eral insects, namely, Coleoptera (Leptinotarsa decemli- neata; de Kort and Koopmanschap, 1987), Isoptera (Reticulitermes flavipes; Okot-Kotber and Prestwich, 1991), Diptera (Sarcophaga bullata; Van Mellaert et al., 1985; Drosophila melanogaster; Shemshedini and Wil- son, 1988; Chironomus thummi; Wisniewski and Streuer- nagel, 1990), Hymenoptera (Apis mellifera; de Kort and Koopmanschap, 1986), and Dictyoptera (Periplaneta americana; de Kort and Koopmanschap, 1986; Leuco- phaea maderae; Rayne and Koeppe, 1988; Nauphoeta cinerea; Kindle et al., 1989; Diploptera punctata; King and Tobe, 1988). Structural studies of lipophorin indicate that the holoprotein consists of two apoproteins, apolipo- protein-I (apoLp-I; M r  250 kDa) and apolipoprotein-II (apoLp-II; M r  85 kDa) (for reviews see Kanost et al.,