Carotenoids undergo a wide range of photochemical reactions in animal, plant, and microbial systems. In photosynthetic organisms, in addition to light harvesting, they perform an essential role in protecting against light-induced damage by quenching singlet oxygen, superoxide anion radicals, or triplet-state chlorophyll. We have determined the crystal structure of a water-soluble orange carotenoid protein (OCP) isolated from the cyanobacterium Arthrospira maxima at a resolution of 2.1 A. OCP forms a homodimer with one carotenoid molecule per monomer. The carotenoid binding site is lined by a striking number of methionine residues. The structure reveals several possible ways in which the protein environment influences the spectral properties of the pigment and provides insight into how the OCP carries out its putative functions in photoprotection.

Singlet oxygen, 1 O 2 , is a highly reactive electronically excited state of oxygen invoked in many physiological and pathological processes. 1 While its generation in biological systems is mainly traced back to photosensitization by sunlight-absorbing cofactors and dark enzymatic pathways, 1 O 2 can also form by photosensitization with aromatic amino acids such as tryptophan (Trp), tyrosine (Tyr), and phenylalanine (Phe), which are abundant light absorbers in the UV-B range (290-320 nm). 2 Notably, though the role of 1 O 2 in the formation of H 2 O 2 by in vitro UV irradiation of aromatic amino acids in immunoglobulins was reported by Wentworth et al., 3 the generation of 1 O 2 by aromatic amino acids and biological macromolecules has not been analyzed quantitatively. 4 In this manuscript we report the quantum yields of 1 O 2 generation upon excitation of Trp, Tyr, and Phe in their zwitterionic forms, as methyl esters, and within a few test proteins and immunoglobulins.The sensitized generation of 1 O 2 starts by excitation of the amino acid by absorption of UV light, followed by intersystem crossing to the triplet state (eqs 1 and 2), and is completed by energy transfer to oxygen in its triplet ground state (eq 3). The quantum yields of 1 O 2 can be obtained in air-saturated solutions by measuring the quantum yield of its near IR emission (eq 4 and Figure 1).The amino acids Phe, Trp, and Tyr, the N-acetylated amino acids NAc-Phe, NAc-Trp, and NAc-Tyr, the proteins bovine serum albumin (BSA) and ovoalbumin (OVA), and the immunoglobulins bovine-IgG, human-IgG, and sheep-IgG were dissolved in either D 2 O or MeCN. Samples were irradiated with a 266 nm pulse from a frequency quadrupled continuum Nd:YAG laser (8 ns, 3 mJ).The near-IR luminescence from O 2 ( 1 ∆ g ) was collected at right angles to excitation after focusing onto the variable slit of a SPEX 1681 monochromator and detected using a Hamamatsu PMT sensitive in the near-IR region. The near-IR luminescence spectrum of singlet oxygen photosensitized by phenylalanine in an air-saturated D 2 O solution is shown in Figure 1. There is a strong peak at about 1270 nm, characteristic of singlet oxygen phosphorescence. A time-resolved decay trace of the observed phosphorescence has a lifetime (τ ∆ ) of about 48 µs, consistent with reported values of 1 O 2 in D 2 O. Solutions of tryptophan, tyrosine, bovine serum albumin (BSA), ovalbumin (OVA), the immunoglobulins bovine IgG, human IgG, and sheep IgG in D 2 O, and the N-acetyl amino acids in MeCN were irradiated under similar conditions to emit spectra consistent with singlet oxygen luminescence. Argon-purged solutions of each compound showed no near-IR luminescence.The quantum yields of oxygen (Φ ∆ ) were determined using methylene blue in D 2 O (Φ ∆ ) 0.52) or MeCN (Φ ∆ ) 0.52) as a reference. 5 Triplicate kinetic analysis of decay traces for Tyr and Trp gave lifetimes for singlet oxygen of 35-40 µs in D 2 O, which are about 20% shorter than those observed with Phe. This may be attributed to quenching, in agreeme...